Method and apparatus for enhancement of transdermal transport

ABSTRACT

According to the present invention, a method for enhancing transdermal transport is disclosed. The method includes the steps of increasing a permeability of an area of a membrane with a permeabilizing device. The membrane may be, inter alia, biologic skin or synthetic skin. The permeabilizing device may be an ultrasound-producing device. A substance is transported into and through the area of the membrane. The substance may be a drug, a vaccine, or a component of interstitial fluid.

BACKGROUND OF THE INVENTION

[0001] 1. Field of the Invention

[0002] This invention relates to transdermal molecular transportation.More specifically, this invention relates to methods and apparatus forthe regulation of skin permeabilization and analysis of analytes inextracted body fluid.

[0003] 2. Description of the Related Art

[0004] Drugs are routinely administered either orally or by injection.The effectiveness of most drugs relies on achieving a certainconcentration in the bloodstream. Although some drugs have inherent sideeffects which cannot be eliminated in any dosage form, many drugsexhibit undesirable behaviors that are specifically related to aparticular route of administration. For example, drugs may be degradedin the GI tract by the low gastric pH, local enzymes or interaction withfood or drink within the stomach. The drug or disease itself mayforestall or compromise drug absorption because of vomiting or diarrhea.If a drug entity survives its trip through the GI tract, it may facerapid metabolism to pharmacologically inactive forms by the liver, thefirst-pass effect. Sometimes the drug itself has inherent undesirableattributes such as a short half-life or a narrow therapeutic blood levelrange.

[0005] Recently, efforts aimed at eliminating some of the problems oftraditional dosage forms involve transdermal delivery of the drugs(TDD). Topical application has been used for a very long time, mostly inthe treatment of localized skin diseases. Local treatments, however,only require that the drug permeate the outer layers of the skin totreat the diseased state, with little or no systemic accumulation.Transdermal delivery systems are designed for, inter alia obtainingsystemic blood levels, and topical drug application. For purposes ofthis application, the word “transdermal” is used as a generic term todescribe the passage of substances into, out of, to, and through theskin.

[0006] TDD offers several advantages over traditional delivery methods,including injections and oral delivery. When compared to oral delivery,TDD avoids gastrointestinal drug metabolism, reduces first-pass effects,and provides sustained release of drugs for up to seven days, asreported by Elias in Percutaneous Absorption:Mechanisms-Methodology-Drug Delivery, Bronaugh, R. L. Maibach, H. 1.(Ed), pp 1-12, Marcel Dekker, New York, 1989.

[0007] The transport of drugs to, into, out of, and through the skin iscomplex since many factors influence their permeation. These include theskin structure and its properties, the penetrating molecule and itsphysical-chemical relationship to the skin and the delivery matrix, andthe combination of the skin, the penetrant, and the delivery system as awhole. Particularly, the skin is a complex structure. There are at leastfour distinct layers of tissue the nonviable epidermis (stratum corneum,SC) the viable epidermis, the viable dermis, the subcutaneous connectivetissue. Located within these layers are the skin's circulatory system,the arterial plexus, and appendages, including hair follicles, sebaceousglands, and sweat glands. The circulatory system lies in the dermis andtissues below the dermis. The capillaries do not actually enter theepidermal tissue but come within 150 to 200 microns of the outer surfaceof the skin.

[0008] In comparison to injections, TDD can reduce or eliminate theassociated pain and the possibility of infection. Theoretically, thetransdermal route of drug administration could be advantageous in thedelivery of many therapeutic drugs, including proteins, because manydrugs, including proteins, are susceptible to gastrointestinaldegradation and exhibit poor gastrointestinal uptake. Proteins, such asinterferon, are cleared rapidly from the blood and need to be deliveredat a sustained rate in order to maintain their blood concentration at ahigh value. Transdermal devices are also easier to use than injections.

[0009] In spite of these advantages, very few drugs and no proteins orpeptides are currently administered transdermally for clinicalapplications because of the low skin permeability to drugs. This lowpermeability is attributed to the SC, the outermost skin layer whichconsists of flat, dead cells filled with keratin fibers (keratinocytes)surrounded by lipid bilayers. The highly-ordered structure of the lipidbilayers confers an impermeable character to the SC (Flynn, G. L., inPercutaneous Absorption: Mechanisms-Methodology-Drug Delivery; Bronaugh,R. L., Maibach, H. 1. (Ed), pages 27-53, Marcel Dekker, New York 1989).Several methods have been proposed to enhance transdermal drugtransport, including the use of chemical enhancers, i.e., the use ofchemicals to either modify the skin structure or to increase the drugconcentration in a transdermal patch (Burnette, R. R., in DevelopmentalIssues and Research Initiatives; Hadgraft J., Guy, R. H., Eds., MarcelDekker 1989; pp. 247-288; Junginger, et al. in Drug PermeationEnhancement; Hsieh, D. S., Eds., pp. 59-90; Marcel Dekker, Inc. New York1994) and the use of applications of electric fields to create transienttransport pathways (electroporation) or to increase the mobility ofcharged drugs through the skin (iontophoresis) (Prausnitz Proc. Natl.Acad. Sci. USA 90, 10504-10508 (1993); Walters, K. A., in TransdermalDrug Delivery: Developmental Issues and Research Initiatives, Ed.Hadgraft J., Guy, R. H., Marcel. Dekker, 1989). Another approach thathas been explored is the application of ultrasound.

[0010] Ultrasound has been shown to enhance transdermal transport ofdrugs across human skin, a phenomenon referred to as sonophoresis (Levy,J. Clin. Invest. 1989, 83, 2974-2078; Kost and Langer in “Topical drugBioavailability, Bioequivalence, and Penetration”; pp. 91-103, Shah V.P., Maibach H. I., Eds. (Plenum: New York, 1993). For example, U.S. Pat.No. 4,309,989 to Fahim and U.S. Pat. No. 4,767,402 issued to Kost et al.both describe the use of ultrasound in conjunction with transdermal drugdelivery. U.S. Pat. No. 4,309,989 discloses the topical application of amedication using ultrasound with a coupling agent such as oil.Ultrasound at a frequency of at least 1000 kHz and a power of one tothree W/cm² was used to create selective localized intracellularconcentration of a zinc-containing compound for the treatment of herpessimplex virus.

[0011] U.S. Pat. No. 4,309,989, the disclosure of which is incorporatedby reference in its entirety, discloses the use of ultrasound forenhancing and controlling transdermal permeation of a molecule,including drugs, antigens, vitamins, inorganic and organic compounds,and various combinations of these substances, through the skin and intothe circulatory system. Ultrasound having a frequency of about 20 kHzand having an intensity between about 0 and 3 W/cm² is used essentiallyto drive molecules through the skin and into the circulatory system.

[0012] Although a variety of ultrasound conditions have been used forsonophoresis, the most commonly used conditions correspond totherapeutic ultrasound (frequency in the range of between one MHz andthree MHz, and intensity in the range of between above zero and twoW/cm²) (such as that described in the Kost et al. patent). It is acommon observation that the typical enhancement induced by therapeuticultrasound is less than ten-fold. In many cases, no enhancement oftransdermal drug transport has been observed upon ultrasoundapplication. Accordingly, a better selection of ultrasound techniques isneeded to induce a higher enhancement of transdermal drug transport bysonophoresis.

[0013] Application of low-frequency ultrasound (between about 20 and 200kHz) can dramatically enhance transdermal transport of drugs, as well asthe extraction and measurement of analyte, as described inPCT/US96/12244 by Massachusetts Institute of Technology. Transdermaltransport enhancement induced by low-frequency ultrasound was found tobe as much as 1000-fold higher than that induced by therapeuticultrasound. Another advantage of low-frequency sonophoresis as comparedto therapeutic ultrasound is that the former can induce transdermaltransport of drugs which do not passively permeate across the skin.

[0014] Ultrasound gels may be used as couplings in most medicalapplications of ultrasound energy. Use of these gels may be messy andlabor-intensive. To overcome problems associated with applyingultrasound with gels and other coupling agents, patches containing therequired components have been developed. A patch adheres to a clean areaof the skin, and drug molecules are continually absorbed through theskin into the bloodstream for systematic distribution. These patchesinclude a drug-containing layer provided near an ultrasonic oscillator.Drug absorption is ensured by the action of the ultrasonic waves fromthe oscillator. The amount of drug released may be controlled by varyingthe ultrasonic wave output from the oscillator, as described in U.S.Pat. No. 5,007,438 to Tachibana, et al., the disclosure of which isincorporated by reference in its entirety. U.S. Pat. No. 4,821,740 toTachibana et al. discloses a kit for providing external medicines thatincludes a drug containing layer and an ultrasonic oscillator forreleasing the drugs for uptake through the surface of the skin. Thetransducer may be battery powered. The application of the ultrasoundcauses the medication to move from the device to the skin and then theultrasound energy may be varied to control the rate of administrationthrough the skin.

[0015] U.S. Pat. No. 5,421,816 to Lipkovker describes ultrasonic energythat releases a stored drug and forcibly moves the drug through the skinof an organism and to the blood stream. A housing includes a cavitydefined by an assembly of ultrasonic transducers and separated from theskin by a polymeric membrane that stores the drug to be delivered. Theultrasonic transducer assembly includes a flat, circular ultrasonictransducer that defines the top of a truncated cone and a polarity oftransducer segments that define the walls of the cone. The resonantfrequency of the planar transducer is lower than the resonant frequencyof the transducer segments. The planar, flat, circular transducergenerates a fixed frequency in the 5 kHz to 1 MHz range, and ultrasonicstimuli impulses for a predetermined period of time, such as 10-20seconds.

[0016] Between the stimuli pulse periods, the transducer segmentsreceive variable frequency ultrasonic pumping pulses. The variablefrequency ultrasonic pumping pulses lie in the 50 MHz to 300 MHz range.The variable frequency ultrasonic pumping pulses are applied to opposingtransducer segments. The transducer segments create beams that impingeon the skin at an oblique angle to create a pulsating wave. Further, thevariable frequency ultrasonic pumping pulses are applied to opposingtransducer segments in a rotating manner to create pulsating waves inthe skin in a variety of directions. The stimuli pulses cause the planartransducer to produce an ultrasonic wave that excites the local nervesin a way that trauma, such as heat and force, excites local nerves. Thevariable frequency ultrasonic pumping pulses cause the transducersegments to produce ultrasonic waves in both the polymeric membrane andthe skin. The ultrasonic waves pump the drug to the polymeric membraneand, then, through skin openings into the underlying blood vessels.

[0017] Thus ultrasound energy may serve to enhance the flux of activepermeate molecules through the skin and other biological membranes byproviding an active energy source, in addition to passive diffusion, topush or pump molecules through pores and channels.

[0018] In addition to there being a need to deliver drugs through theskin, there is a major medical need to extract analytes through theskin. For example, it is desirable for diabetics to measure bloodglucose several times per day in order to optimize insulin treatment andthereby reduce the severe long-term complications of the disease.Currently, diabetics do this by pricking the highly vascularizedfingertips with a lancet to perforate the skin, then milking the skinwith manual pressure to produce a drop of blood, which is then assayedfor glucose using a disposable diagnostic strip and a meter into whichthis strip fits. This method of glucose measurement has the majordisadvantage that it is painful, so diabetics do not like to obtain aglucose measurement as often as is medically indicated.

[0019] Therefore, many groups are working on non-invasive, and lessinvasive means to measure glucose, such as micro lancets that are verysmall in diameter, very sharp, and penetrate only to the interstitium(not to the blood vessels of the dermis). A small sample, from about 0.1to two μl, of interstitial fluid is obtained through capillary forcesfor glucose measurements. Other groups have used a laser to breach theintegrity of the stratum corneum and thereby make it possible for bloodor interstitial fluid to diffuse out of such a hole or to be obtainedthrough such a hole using pneumatic force (suction) or other techniques.An example of such a laser based sampling device is disclosed in U.S.Pat. No. 5,165,418 to Tankovich and WPI ACC No: 94-167045/20 by Budnik(assigned to Venisect, Inc.).

[0020] A problem with methods that penetrate the skin to obtaininterstitial fluid is that interstitial fluid occurs in the body in agel like form with little free fluid and, in fact, is even undernegative pressure that limits the amount of free interstitial fluid thatcan be obtained. When a very small hole is made in the skin, penetratingto a depth such that interstitial fluid is available, it takes a greatdeal of mechanical force (milking, vacuum, or other force) to obtain therequisite quantity of blood, or interstitial fluid, used in a glucosemeter.

[0021] Channeling of ultrasound geometrically is one way to applyultrasound to a small area. Channeling of ultrasound is disclosed in PCTPatent Application No. PCT/US97/11559 entitled “Ultrasound Enhancementof Transdermal Transport” by Sontra L. P. et al., filed Jun. 30, 1997,and incorporated by reference in its entirety. The oscillation of asmall element near or in contact with the surface of the skin is anotherway to apply ultrasound to a small area. Large forces can be producedlocally, resulting in cavitation, mechanical oscillations in the skinitself, and large localized shearing forces near the surface of theskin. The element can also produce acoustic streaming, which refers tothe large convective flows produced by ultrasound. This appears to aidin obtaining a sample of blood or interstitial fluid without having to“milk” the puncture site. Ultrasound transducers are known to rapidlyheat under continuous operation, reaching temperatures that can causeskin damage. Heat damage to the skin can be minimized by using atransducer that is located away from the skin to oscillate a smallelement near the skin. In the case of analyte extraction, compoundspresent on the surface of and/or in the skin can contaminate theextracted sample. The level of contamination increases as skin surfacearea increases. Surface contamination can be minimized by minimizing thesurface area of ultrasound application. Thus, skin permeability can beincreased locally, and transiently through the use of the methods anddevices described herein, for either drug delivery or measurement ofanalyte.

[0022] Moreover, it has been disclosed that the application ofultrasound is only required once for multiple deliveries or extractionsover an extended period of time rather than prior to each extraction ordelivery. That is, it has been shown that if ultrasound having aparticular frequency and a particular intensity is applied, multipleanalyte extractions or drug deliveries may be performed over an extendedperiod of time. For example, if ultrasound having a frequency of 20 kHzand an intensity of about 10 W/cm² is applied, the skin retains anincreased permeability for a period of up to four hours. This isdescribed more particularly in U.S. patent application Ser. No.09/227,623 entitled “Sonophoretic Enhanced Transdermal Transport” byMitragotri et al., filed on Jan. 8, 1999, and in PCT Application No.PCT/US99/00437 entitled “Sonophoretic Enhanced Transdermal Transport” bySontra Medical et al., filed Jan. 8, 1999 the disclosures of which arehereby incorporated by reference in their entireties.

[0023] Nevertheless, the amount (e.g., duration, intensity, duty cycleetc.) of ultrasound necessary to achieve this permeability enhancementvaries widely. Several factors of the nature of skin must be considered.For example, the type of skin which the substance is to pass throughvaries from species to species, varies according to age (e.g., the skinof an infant has a greater permeability than that of an older adult),varies according to local composition, thickness and density, varies asa function of injury or exposure to agents such as organic solvents orsurfactants, and varies as a function of some diseases, such aspsoriasis, or abrasion. Moreover, as discussed above, overexposure toultrasound and cavitation can cause damage to the skin through heatingand increased pressure. Therefore, it is necessary to control theultrasound application in order to enable clinically useful transdermaltransport.

SUMMARY OF THE INVENTION

[0024] Therefore, a need has arisen for a method and apparatus forregulation of skin permeabilization through a feedback system.

[0025] A need has arisen for a method and apparatus that providescontrolled enhancement of transdermal transport.

[0026] A need has also arisen for a system and method for extraction andanalysis of at least one analyte in a body fluid.

[0027] A need has also arisen for a method and apparatus forsonophoretic drug delivery.

[0028] According to the present invention, a method for enhancingtransdermal transport is disclosed. The method includes the steps ofincreasing a permeability of an area of a membrane with a permeabilizingdevice. Next, the permeability of the area of membrane is monitored. Asubstance is transported into and through the area of the membrane.

[0029] According to one embodiment of the present invention, a methodfor enhancing permeability of an area of skin is disclosed. The methodcomprises applying ultrasound to the area of skin. While the ultrasoundis being applied, electricity (e.g., an ac current source or an acvoltage source) is applied to the area of skin. While the electricity isbeing applied to the area of skin, a first electrical parameter of thearea of skin is measured. Based on the measured first electricalparameter, the ultrasound is controlled.

[0030] According to another embodiment, the present invention comprisesan apparatus for enhancing the permeability of an area of skin. Theapparatus includes an ultrasound-producing device configured to applyultrasound to the area of skin, an electrical source operable to applyelectricity to the area of skin, a circuit to measure a first electricalparameter of the area of skin, and a controller responsive to thecircuit and operable to control the ultrasound-producing device.

[0031] According to another embodiment, the present invention comprisesa method for enhancing the permeability of an area of skin. The methodbegins by creating a volume of fluid adjacent the area of skin. Thefluid has an initial concentration of a first substance. Ultrasound isthen applied to the area of skin. While the ultrasound is being applied,changes in the concentration of the first substance are monitored.Finally, the method controls the ultrasound based on the changes in theconcentration of the substance.

[0032] According to another embodiment, the present invention comprisesa method for enhancing the permeability of an area of skin. The methodbegins by creating a volume of fluid adjacent the area of skin whosepermeability is to be enhanced. A reference value for an electricalparameter of the volume of fluid is then determined. The method thenapplies ultrasound to the area of skin and monitors changes in theelectrical parameter of the volume of fluid. Finally, the ultrasound iscontrolled based on the changes in the electrical parameter of thevolume of fluid.

[0033] According to another embodiment of the present invention, amethod for regulating skin permeabilization is disclosed. The methodcomprises coupling a first electrode in electrical contact with a firstarea of skin. A second electrode is placed in electrical contact with asecond area of skin. The initial conductivity between these sites ismeasured, and then a skin permeabilizing method, such as ultrasound, isapplied to the first area of skin. The conductivity between the firstarea and second area is measured again. Mathematical analysis or signalprocessing is performed on the conductivity information. Next,parameters describing the kinetics of skin conductance are calculated.Next, once the desired value of the parameters are reached, the skinpermeabilizing step is terminated.

[0034] According to another embodiment, the present invention comprisesan apparatus for enhancing permeability of an area of skin. Theapparatus includes a first electrode for coupling in electrical contactwith a first area of skin, and a second electrode for placement inelectrical contact with a second area of skin. A skin permeabilizingdevice, such as an ultrasound-producing device, is provided to apply askin permeabilizing treatment to the skin at the first area. A means formeasuring the conductivity between the first area and second area areprovided. A controller, for performing mathematical analysis or signalprocessing on the conductivity information, and for calculating thekinetics of skin conductance is provided. The controller also controlsthe skin permeabilizing device.

[0035] According to another embodiment of the present invention, amethod for extraction and analysis of at least one analyte in a bodyfluid is disclosed. According to this method, first the permeabilitylevel of an area of skin is increased. Next, a body fluid is extractedfrom the area of skin. Then, the body fluid is collected. Next, adetermination is made as to the presence of at least one analyte in thebody fluid.

[0036] The body fluid may be extracted by physical forces, chemicalforces, biological forces, vacuum pressure, electrical forces, osmoticforces, diffusion forces, electromagnetic forces, ultrasound forces,cavitation forces, mechanical forces, thermal forces, capillary forces,fluid circulation across the skin, electro-acoustic forces, magneticforces, magneto-hydrodynamic forces, acoustic forces, convectivedispersion, photo acoustic forces, by rinsing body fluid off skin, andany combination thereof The body fluid may be collected by a collectionmethod including absorption, adsorption, phase separation, mechanical,electrical, chemically induced, and a combination thereof. The presenceof an analyte may be sensed by a sensing method includingelectrochemical, optical, acoustical, biological, enzymatic technology,and combinations thereof.

[0037] According to another embodiment of the present invention, asystem for extraction and analysis of at least one analyte in a bodyfluid is disclosed. The system comprises a transducer for increasing thepermeability of an area of skin; an extraction device for extractinginterstitial fluid from the area of skin; a collection device forcollecting the extracted interstitial fluid; and a sensing device forsensing the presence of at least one analyte in the extractedinterstitial fluid.

[0038] According to another embodiment of the present invention, amethod for blood glucose determination is disclosed. The method includesfirst increasing a permeability of an area of skin. Next, interstitialfluid, or components thereof, is extracted from the area of skin. Inanother embodiment, the interstitial fluid, or components thereof,diffuse through the skin, and are collected. Next, the interstitialfluid is collected in a gel. The gel may contain at least one glucosesensitive reagent that changes at least one characteristic of the gel,such as color, when glucose is present. Finally, the change in thecharacteristic of the gel is monitored.

[0039] According to another embodiment of the present invention, asystem for blood glucose determination is disclosed. The systemcomprises a transducer for increasing the permeability of the skin; anextraction device for extracting interstitial fluid from the skin; acollection device for collecting the extracted interstitial fluid; a gelhaving at least one glucose sensitive reagent that changes acharacteristic of the gel when glucose is present; and a monitoringdevice for monitoring a change in the characteristic of said gel.

[0040] According to another embodiment of the present invention, a drugdelivery patch apparatus is disclosed. The apparatus includes anultrasound transducer for applying ultrasound to a membrane. Themembrane may include biological membranes, synthetic membranes, or acell culture. A biological membrane may include skin, mucosal and buccalmembranes. The apparatus further includes a power source coupled to thetransducer. The apparatus further includes drug molecules between thetransducer and the membrane, and an attaching device that attaches theapparatus to the membrane. According to another embodiment, theapparatus further includes drive electronics coupled to the transducersuch that the drive electronics enables the transducer to applyultrasound. According to another embodiment, the apparatus furtherincludes an interface coupled to the drive electronics.

[0041] The drug delivery patch apparatus may include the interface,drive electronics, power source, transducer, drug molecules, andattaching device contained within the patch for transdermal deliverythrough the membrane. Alternatively, the transducer and the drugmolecules as well as the attaching device may be contained in the patch.The power source and interface may be connected to the patch with aconnecting wire, or without a wire. Alternatively, the drug deliverypatch may include the power source, the transducer, the drug moleculesand the attaching device within the patch. The interface may be locatedelsewhere and communicates to the patch through hardwires, infrared,fiber optics, or telemetry.

[0042] According to another embodiment of the present invention, amethod for transdermal vaccination by sonophoresis is disclosed.According to the one embodiment, the method comprises the steps ofenhancing the permeability of the skin by the application of ultrasound;providing a vaccine to the permeabilized skin, and delivering thevaccine to the skin cells, for example, Langerhans cells, dendric cells,and keratinocytes.

[0043] In another embodiment of the present invention, ultrasound isused to enhance the permeability of the skin. In another embodiment ofthe present invention, the steps of increasing the permeability of theskin and providing a vaccine to the permeabilized skin occursimultaneously.

[0044] In another embodiment of the present invention, ultrasound isused to irritate or inflame an area of skin. Next, a vaccine is providedto the irritated or inflamed skin. This is more effective in inducingthe immune response of the body.

BRIEF DESCRIPTION OF THE DRAWINGS

[0045] The features and objects of the present invention, and the mannerof attaining them is explained in detail in the following DETAILEDDESCRIPTION OF THE PREFERRED EMBODIMENTS of the invention when taken inconjunction with the accompanying drawings wherein:

[0046]FIG. 1 depicts a schematic of an electrical model for skin,

[0047]FIG. 2 depicts a flow chart of a method for controlled enhancementof transdermal delivery according to one embodiment of the presentinvention;

[0048]FIG. 3 depicts a diagram of a circuit that enhances skinpermeability and monitors enhancement of skin permeability according toone embodiment of the present invention;

[0049]FIG. 4 depicts a permeability monitoring circuit according toanother embodiment of the present invention;

[0050]FIG. 5 depicts a permeability monitoring circuit according to oneembodiment of the present invention;

[0051]FIG. 6 depicts a flow chart of a method for controlled enhancementof transdermal delivery according to one embodiment of the presentinvention;

[0052]FIG. 7 depicts a flow chart of a method for controlled enhancementof transdermal delivery according to one embodiment of the presentinvention;

[0053]FIG. 8 depicts a flow chart of a method for controlled enhancementof transdermal delivery according to one embodiment of the presentinvention;

[0054]FIG. 9 depicts the time variation of the skin conductance whilebeing exposed to ultrasound according to an example;

[0055]FIG. 10 shows a relationship between the inflection time and thetime to pain on various volunteers according to an example;

[0056]FIG. 11 depicts a flowchart of a method of determining when toterminate the application of ultrasound;

[0057]FIG. 12 depicts example graphs of the method of FIG. 11;

[0058]FIG. 13 depicts a flowchart of a method for extraction andanalysis of at least one analyte in a body fluid according to oneembodiment of the present invention;

[0059]FIG. 14 depicts a drawing of a tensioner according to oneembodiment of the present invention;

[0060]FIG. 15 depicts a flowchart of a method of determination of bloodglucose according to one embodiment of the present invention;

[0061]FIG. 16 illustrates a drug delivery patch apparatus in accordancewith one embodiment of the present invention;

[0062]FIG. 17 illustrates a cross-sectional view of a transducer inaccordance with one embodiment of the present invention;

[0063]FIG. 18 illustrates a drug delivery patch apparatus having afeedback mechanism in accordance with one embodiment of the presentinvention, and

[0064]FIG. 19 depicts a flowchart of the method for transdermalvaccination by sonophoresis according to one embodiment of the presentinvention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0065] As used herein, the terms skin permeabilizing method or deviceincludes the application of ultrasound, chemicals, electroporation,mechanical, disrupting devices, tape-stripping, and laser, and devicesfor the application of the same. In addition, the term skin includesmembranes, such as biologic and synthetic skin.

[0066] Ultrasound is generally defined as sound at a frequency ofgreater than about 20 kHz. Therapeutic ultrasound is typically between20 kHz and 5 MHz. Sonophoresis is defined as the application ofultrasound to the skin resulting in enhanced transdermal transport ofmolecules. Low frequency sonophoresis or ultrasound is defined assonophoresis or ultrasound at a frequency that is less than 2.5 MHz,more typically less than 1 MHz, more preferably in the range of 20 to100 kHz.

[0067] Near ultrasound is typically about 10 kHz to 20 kHz. It should beunderstood that in addition to ultrasound, near ultrasound may also beused in the embodiments of the present invention.

[0068] 1. Enhancement and Regulation of Skin Permeability

[0069] The use of ultrasound to facilitate transdermal transport isknown. The mechanism by which ultrasound is used to facilitatetransdermal transport has differed. In the context of transdermaldelivery systems, ultrasound was initially used as a driving force thatessentially pushed drugs through the skin and into the circulatorysystem. Ultrasound is also used to increase the permeability of theskin. That is, application of ultrasound having a particular frequencywill disorganize the lipid bilayer in the skin and thus increase thepermeability of the skin. In this context, either drugs can be deliveredthrough the skin to the body or analyte or analytes can be extractedthrough the skin from the body. A driving force of some type is stillrequired, but the required intensity of the driving force is decreased.For example, a concentration gradient is generally sufficient drivingforce for transdermal transport through skin whose permeability has beenenhanced using ultrasound.

[0070] Regardless of which mechanism is used, the ultrasound still needsto be controlled. That is, overexposure to ultrasound may cause skindamage from increased heat, increased pressure and other factors.Therefore according to various embodiments of the present invention, amethod and an apparatus for controlled enhancement of skin permeabilityare disclosed. The method and apparatus, according to the presentinvention, focus on the use of electrical parameters of the skin as aproxy for skin permeability. The skin can be modeled using an R-Ccircuit similar to that shown in FIG. 1. The “skin circuit,” shown inFIG. 1, consists of a resistor R₁ in parallel with a capacitor C, bothof which are in series with a resistor R₂. For normal, intact skin, ofan area of about 1.7 cm², the value for R₁ is about 100 kΩ, the valuefor C is about 13 μF and the value for R₂ is about 2 kΩ. Of course,these values will vary from person to person depending on skin type andcondition. By its nature, the behavior (i.e., the frequency response) ofthe “skin circuit” changes in response to excitations having differentfrequencies. For example, under normal conditions, the impedance of thiscircuit will decline sharply as frequency increases, for example, from10 Hz to 1 kHz. That is, at low frequencies, the capacitive component tothe impedance of the parallel combination of R₁ and C is significant andtherefore the overall impedance of the circuit is high. At higherfrequencies, however, the capacitive component to the impedance of theparallel combination decreases and, therefore, the overall impedance ofthe “skin circuit” declines.

[0071] Various electrical parameters of the skin (e.g., impedance,conductance, inductance and capacitance) have values that correlate withskin permeability. For example, in the circuit of FIG. 1, the value ofR₁ significantly decreases as the skin becomes permeable. For example,R₁ may drop to a value around 5 kΩ for a skin area of about 1.7 cm².Therefore, the frequency response of the overall skin circuit becomesmuch flatter as frequency increases. That is, the difference between theimpedance of the circuit at 10 Hz and 1 kHz would not be nearly assignificant as at 10 Hz alone. Thus, the methods and apparatus of thepresent invention measure one or more electrical parameters of an areaof skin that is being exposed to ultrasound and then adjust the sourceof ultrasound based on the measured parameters.

[0072] According to one embodiment of the present invention, a methodfor controlled enhancement of skin permeability is disclosed, and willbe explained in conjunction with FIG. 2. Typically, when a skinpermeabilizing device, such as an ultrasonic device, is used to enhancetransdermal transport properties, the skin permeabilizing device isapplied to a small patch of skin. In step 202, a baseline measurementfor some electrical parameter is determined for the patch of skin towhich the skin permeabilizing will be applied to determine baselineparameters. In one embodiment, a baseline impedance is measured for thepatch of skin to which the skin permeabilization device is to beapplied. In other embodiments, a baseline conductance, a baselinecapacitance, a baseline inductance, or a baseline capacitance may bemeasured.

[0073] The baseline measurement may be made by using two or moreelectrodes. As is shown in greater detail in FIG. 3, an electrode, suchas source electrode 310, is coupled to the patch of skin to whichultrasound is to be applied. Source electrode 310 does not have to makedirect contact with the skin. Rather, it may be electrically coupled tothe skin through the medium that is being used to transmit ultrasound. Asecond or counter electrode, such as conductive band 312, may bepositioned on a second area of skin that the skin permeabilizing devicewill not be applied to. This second area of skin can be adjacent to thepatch of skin to which the skin permeabilizing device will be applied,or it can be distant from that patch of skin.

[0074] In one embodiment, the ultrasonic transducer and horn that applythe ultrasound double as the source electrode through which electricalparameters of the patch of skin may be measured, and is coupled to theskin through a conductive solution, such as saline, used as anultrasound medium. In another embodiment, a separate electrode may beaffixed to the area of skin that ultrasound will be applied to and isused as the source electrode. In still another embodiment, the housingof the device used to apply ultrasound to the area of skin may be usedas the source electrode. The electrode can be made of any suitableconducting material including, for example, metals and conductingpolymers.

[0075] In order to achieve an accurate electrical reading, the counterelectrode should make sufficient contact with the skin. This can beachieved in a number of ways. In one embodiment, the counter electrodeis applied directly to the epidermis of the skin. That is, the counterelectrode is applied to an area of skin from which the stratum corneumhas been removed. The stratum corneum may be removed in a number ofways. According to one embodiment, the stratum corneum is removed bytape stripping. In one embodiment, sufficient electrical contact betweenthe skin and the counter electrode is created by using a counterelectrode having a large surface area. More specifically, a conductivepolymeric path or metallic foil patch having an area much larger thanthe skin area exposed to ultrasound is used. The large area of thecounter electrode in this embodiment decreases its impedance and allowsaccurate measurements of the electrical parameter of the area of skinexposed to ultrasound. In one specific embodiment a conductive band iswrapped around the subject's arm and used as the counter electrode. Inanother embodiment, the counter electrode may be placed in a handle ofthe skin permeabilizing device, to which a subject grasps duringoperation.

[0076] In another embodiment, the counter electrode surrounds the skinpermeabilizing device.

[0077] When the two electrodes are properly positioned, the baselinemeasurement may be made by applying an electrical signal to the patch ofskin through the electrodes. The electrical signal supplied preferablyhas a sufficient intensity so that the electrical parameter of the skincan be measured, but a suitably low intensity so that the electricalsignal does not cause damage to the skin or any significantelectrophoresis effect for the substance being delivered. In oneembodiment, a 10 Hz AC source may be used to create a voltagedifferential between the source electrode and the counter electrode. Inone embodiment, in order to avoid a risk of permanent damage to theskin, the voltage supplied does not exceed 500 mV, and, preferably, doesnot exceed 100 mV. In another embodiment, an AC current source is used.The current source may also be similarly limited. The baselinemeasurement is made after the source has been applied using appropriatecircuitry. In one embodiment, a resistive sensor is used to measure theimpedance of the patch of skin at 10 Hz. In another embodiment, a 1 kHzsource is used. Sources of other frequencies are also possible.

[0078] Experiments were performed on human volunteers to ensure that theabove described electrode placement would provide accurate measurements.A first glass chamber (˜1.5 cm² in area) was placed on the forearm andwas secured in place with an elastic strap. This first chamber wasfilled with 2 ml of 1% sodium lauryl sulfate (SLS) in saline. Anultrasound horn was place within the chamber and used to applyultrasound to the skin. Additionally, an electrode used to measureelectrical parameters was incorporated into the horn.

[0079] A second small chamber (˜1.5 cm²) was placed on the subject's armin order to measure skin conductivity. This is referred to as thereference chamber. The skin under the reference chamber wastape-stripped using scotch tape to remove the stratum corneum. Thisprocess involved placing a piece of scotch tape (1.5 cm wide and 3 cmlong) on the subject's arm and removing it. This procedure is repeated˜25 times in order to remove the stratum corneum from the designatedarea. An electrode was then placed on the skin under the chamber.

[0080] Another electrode is placed on the subject's arm. This electrode25 consisted of a large piece of aluminum foil placed on intact skin.Ultrasound (27 kHz, ˜10 μm tip displacement, pulsed: 5 sec. on/5 sec.off) was applied to the first chamber. The conductance of the skinexposed to ultrasound was measured with both counter electrodes (tapestripped and intact). The measured conductances were similar thusproving that a large counter electrode placed over intact skin can besuccessfully used to measure skin conductance during sonophoresis.

[0081] Referring again to FIG. 2, in step 204, the skin permeabilizingdevice, such as an ultrasound providing device, is applied to the patchof skin. Although the exact ultrasound parameters are not the subject ofthis invention, according to one embodiment using an ultrasonic deviceas a skin permeabilizing device, ultrasound having a frequency of about20 kHz, and an intensity of about 10 W/cm² may be used to enhance thepermeability of the patch of skin to be used for transdermal transport.

[0082] After the skin permeabilizing device has been turned on, in step206 the permeability of the patch of skin is monitored. Morespecifically, and as discussed above, electrical parameters of the patchof skin are used as a proxy for skin permeability. That is, what isactually being monitored is the electrical parameter for which abaseline measurement was made in step 202. The monitoring measurementsare made using the same electrode set up that was used to make thebaseline measurement.

[0083] In step 208, the skin permeabilizing device is controlled basedon the monitoring measurements made in step 206. In one embodiment, themonitoring measurements are fed back to a microcontroller that is usedto control the skin permeabilizing device. When ultrasound is used, thepermeability enhancement obtained by supplying ultrasound is limited.That is, once a certain permeability is reached, the further applicationof ultrasound will not further enhance skin permeability. Overexposureto ultrasound, or cavitation caused thereby, may result in damage to theskin from localized pressure, temperature increases, and shear stresses.Therefore, in one embodiment, when the parameter being monitored reachesis predetermined value, the ultrasound-producing device is turned off.If the parameter being monitored has not reached the predeterminedvalue, the measurement is repeated until the predetermined value isreached.

[0084] The predetermined value may depend upon a number of factorsincluding, inter alia, the skin characteristics of the individual, thedrug to be delivered or the analyte or analytes to be extracted (becauseof varying molecule sizes), and the frequency of the excitation source.As is apparent to one of ordinary skill in the art, a specificcorrelation between the electrical parameter being used and skinpermeability may be determined by conducting experiments and usingexperimental data. The predetermined value may then be determined on asubject-by-subject basis, taking into account all appropriate factorsand the empirical data.

[0085] According to another embodiment, the intensity of the skinpermeabilizing device may be gradually scaled back as the point ofmaximum permeability enhancement is approached. In one embodiment, asthe parameter being monitored reaches 50% of the predetermined value,either the intensity or the duty cycle may be reduced by a predeterminedamount, such as 50%. This is done so that the predetermined value is not“overshot,” thereby increasing the risk of skin damage. Additionalcontrols are possible. For example, in another embodiment, the intensitymay be scaled back when the parameter being monitored reaches 25%, 50%and 75% of the predetermined value.

[0086] According to another embodiment, permeability enhancement controlmay be accomplished using two electrical sources having differentfrequencies. This method relies on the observation, discussed above,that as the skin becomes more permeable, the frequency response of theskin becomes flatter. In this embodiment, the initial step 202 ofmeasuring a baseline for the parameter is unnecessary because theultrasound control is based on a differential between the parametervalue at two different frequencies of excitation. Nevertheless, abaseline measurement may still be desirable in order to determine therange of values to expect. In this embodiment, the electrode arrangementmay be the same as that described above. And, step 204 of beginningultrasound application is also the same as recited above. Thus, thedetails of these steps will not be reiterated.

[0087] After the skin permeabilizing has begun, in step 206, skinpermeability is monitored. In this embodiment, skin permeability is alsomonitored using an electrical parameter measured from the skin as aproxy. This embodiment differs from the first embodiment in that theelectrical parameter is measured at two frequencies. In one embodiment,the impedance of the skin is measured at frequencies of 10 Hz and 1 kHz.These measurements are then used to control the skin permeabilizingdevice.

[0088] According to this embodiment, in step 208 the parametermeasurement at a first frequency is compared with the parametermeasurement at a second frequency to determine whether the twomeasurements are within a predetermined differential. If the two valuesare within a predetermined differential, it provides an indication thatthe frequency response of the skin has flattened and, therefore, is anindication that the skin has reached an enhanced level of permeability.At this point, the skin permeabilizing device is turned off. In oneparticular embodiment, an impedance of the skin is measured at 10 Hz andat 1 kHz. And, if the two impedance measurements, are within 20% of eachother, the skin permeabilizing device may be turned off.

[0089] The rate of change in the parameter measurements may also be usedto determine a point at which the skin permeabilizing device is scaledback or discontinued. The rate of change of one, or both, or theparameters may be used. In another embodiment, the rate of change of thedifference between the two parameters may also be used. As the rate ofchange reaches a predetermined value, the intensity of the skinpermeabilizing device may be gradually scaled back or discontinued, in amanner similar to that discussed above.

[0090] As discussed above, the predetermined differential value maydepend upon a number of factors, including, inter alia, the skincharacteristics of the individual, the drug to be delivered or theanalyte to be extracted (because of varying molecule sizes), and thefrequencies of the exciting sources. Therefore, the predetermineddifferential is determined on a subject-by-subject basis taking intoaccount all appropriate factors. Empirical data may be used to determinea precise value for the predetermined differential.

[0091] In a modification of this embodiment, the intensity of the skinpermeabilizing device may be gradually scaled back as the point ofmaximum permeability enhancement is approached. For example, as thedifferential between the two parameter measurements approaches 50% ofthe predetermined differential value, either the intensity or the dutycycle may be reduced by a predetermined amount, such as 50%. Additionalcontrols are possible. For example, in another embodiment, the intensityis scaled back when the differential between the two parameters beingmonitored reaches 25%, 50% and 75% of the predetermined differentialvalue.

[0092] In vitro experiments were performed to assess the above twosource method. Pig skin was mounted on diffusion cells. Skin was mountedon the diffusion cell and was exposed to ultrasound using 1% SodiumLauryl Sulfate and saline solution as a coupling medium. Skinconductance was measured by placing two electrodes across the skin. Theimpedances were measured at two frequencies: 10 Hz and 1 kHz. Theimpedances measured at the frequencies differed by about 25 fold priorto application of ultrasound when the skin was not permeable. Uponsonication, the difference between the impedances at two frequenciesdecreased. The decrease in the differential impedance increased withtime. When the skin was highly permeable, the impedances at twofrequencies differed only by ˜20%. Thus the difference between theimpedances measured at two frequencies may be used to determine thelevel of permeabilization and stop sonication.

[0093] The methods described above use a single electrical parameter tocontrol the ultrasound-producing device. Nevertheless, control of theultrasound-producing device may also be based on two or more electricalparameters.

[0094] According to another embodiment of the present invention, anapparatus for controlled enhancement of transdermal transport 300 isdescribed in conjunction with FIG. 3. Apparatus 300 uses anultrasound-producing device as the skin permeabilizing device; it shouldbe noted that other devices for increasing the skin permeability may beused in place of the ultrasound-producing device. For example, thepermeability of the skin may be increased through the application ofelectric fields, chemicals, mechanical forces, needles, and magneticforces.

[0095] Apparatus 300 includes ultrasound transducer/horn combination302, source 304, bandpass filter 306, permeability monitoring circuit308, source electrode 310, return electrode 312, and microcontroller314. Permeability monitoring circuit 308 comprises current sensor 315,amplifier 316, A/D converter 318, and resistor 320.

[0096] Ultrasound transducer/horn combination 302 is used to applyultrasound to the area of skin 322. Transducer 302 may be any knownultrasound transducer, such as a piezoelectric transducer, a ceramictransducer, or polymer block transducer. The horn can have any knownconfiguration. In one embodiment the horn is made of a conductive metal.

[0097] As described above, while the ultrasound is being supplied to thearea of skin, it is important to monitor the skin permeability andcontrol the ultrasound application so that the skin will not beoverexposed to ultrasound. Apparatus 300 may include the electricalcontrol circuitry elements described above in order to accomplish thismonitoring and control. Specifically, source 304 and bandpass filter 306are provided to drive the electrical control circuitry. That is, inorder to obtain the electrical parameter measurements used forcontrolling source 304, a small signal is passed through the area ofskin. In one embodiment of the present invention, source 304 provides a10 Hz AC square wave voltage that is used to monitor the permeability ofthe area of skin in apparatus 300. Bandpass filter 306 is provided toconvert the square wave into a sinusoid.

[0098] Source electrode 310 and return electrode 312 provide anelectrical path through which electrical parameters of the area of skin322 can be measured. Source electrode 310 may be incorporated intotransducer/horn combination 302, and is preferably formed of anysuitable conductive material. In one embodiment, the ultrasound horn ismetal and is used as the source electrode. Return electrode 312 is aconductive band and is preferably formed from a conductive polymericpath or a metallic foil.

[0099] Permeability monitoring circuit 308 comprises circuitry designedto measure an electrical parameter of the skin as a proxy for thepermeability of the skin. More specifically, according to one embodimentof the present invention, permeability monitoring circuit 308 comprisescircuitry designed to measure the current flow through the area of skin322 and to convert that measurement in to a form suitable for use bymicrocontroller 314. Permeability monitoring circuit 308 comprisescurrent sensor 315 that is operable to measure the impedance of area ofskin 322. Current sensor 315 may be any sensor that may be used tomeasure current, and, in one embodiment, current sensor 315 is a 1 kΩcurrent sense resistor where the output voltage generated is 1000 timesthe current flowing through the skin. The output of current sensor 315is an analog signal that should be digitized before it may be used bymicrocontroller 315. Amplifier 316 and resistor 320 serve to amplify theoutput voltage of current sensor 315 so that it may be digitized by A/Dconverter 318. A/D converter 318 may be any suitable A/D converter.

[0100] The signal from A/D converter 316 may then be provided tomicrocontroller 314. Microcontroller 314 may be any suitablemicrocontroller. Microcontroller 314 is programmed to control transducerdriver circuit 324 as described above. In one embodiment,microcontroller 314 determines whether the signal from permeabilitymonitoring circuit 308 is greater than some predetermined value. If so,microcontroller 314 may turn off the ultrasound by, for example,shutting off the D.C. supply for transducer driver circuit 324.Microcontroller 314 may also be configured to provide other controls,such as altering the duty cycle of transducer driver circuit 324 throughthe phase lock loop circuit.

[0101] According to one embodiment of the present invention, additionalcontrols and a user interface may be provided. Fluids controller 330controls the pumps and fluids for the system. Pump 332 may be providedto provide a seal between transducer 302 and the surface of skin 322.Pump 334, in conjunction with valve 336, may be used to fill andevacuate the chamber of transducer 302. The coupling fluid used intransducer 302 may be provided in cartridge 338. Other devices andmethods for providing coupling fluid may also be used.

[0102] A user interface may also be provided. User interface 340includes low battery sensor 342, which may include a comparator. Switch344 may be provided to turn on or off the ultrasound-producing device.Input 346 may be provided to allow a user to adjust the ultrasoundintensity. The ultrasound level may be provided in display 350. Thepermeability level of the skin may be provided in display 352.Indicators 354 and 356 may be provided to alert the user of theoperation of the ultrasound, as well as a when there is a low battery.Additional controls and displays may be provided, as required, toprevent a user from applying ultrasound of a harmful intensity orduration, or to prevent ultrasound from being applied before the systemis ready (i.e., before coupling fluid is provided for transducer 302,etc.).

[0103] The circuitry described above may be replaced with other elementsif the electrical parameter measurements are accomplished in a differentway. More specifically, the circuitry shown in FIGS. 4 or 5 could beused in place of source 304 bandpass filter 306, and permeabilitymonitoring circuit 308 if the control methodology using sources at twofrequencies was to be used. FIG. 4 schematically depicts one embodimentof a circuit useful for implementing dual frequency control of skinpermeability. The circuit comprises sources F₁ and F₂ that supply twodistinct AC signals to the area of skin to which ultrasound is beingapplied. In one embodiment, sources F₁ and F₂ comprise a 10 Hz and a 1kHz current source respectively. These sources are alternately appliedto the area of skin through a microprocessor controlled switch. In theembodiment shown in FIG. 3, microcontroller 314 would control the switchso that sources F₁ and F₂ alternately excite the skin.

[0104] After excitation by one of the sources, the impedance of the skinis measured by measuring the voltage V₁. That is, V₁ is transmitted to amicroprocessor (e.g., microcontroller 314 in FIG. 3) through gaincircuit 402, diode 404, capacitor C₁, and output resistors R₀₁ and R₀₂.The combination of diode 404 and capacitor C₁ comprises an AC to DCconverter suitable for input to an A/D converter to transform the analogsignal from gain circuit 402 to a digital signal suitable for use by amicroprocessor. Output resistors R₀₁ and R₀₂ provide impedance matchingand filtering for the microprocessor, respectively.

[0105] In operation, the circuit of FIG. 4 in conjunction with asuitably programmed microcontroller alternately applies a 10 Hz and a 1kHz AC source to the skin. The circuit, in conjunction with themicroprocessor, measures the impedance of the skin at both frequencies.The microcontroller makes suitable adjustments to theultrasound-producing device based on the differential between theimpedance of the skin at 10 Hz and the impedance of the skin at 1 kHz.

[0106]FIG. 5 schematically depicts yet another embodiment ofpermeability monitoring circuit for use with multiple frequencyexcitation. In the circuit of FIG. 5, sources F₁ and F₂ are appliedsimultaneously through adder circuit 502 to the area of skin to whichultrasound is being applied. The output signal from the skin is then fedto two bandpass filters 504 and 506. Elements C₁, C₂ and R₁ of bandpassfilter 504 are preferably chosen to create a pass band centered aroundthe frequency of source F₁. Elements C₁, C₄ and R₂ of bandpass filter506 are preferably chosen to create a pass band centered around thefrequency of source F₂. The output signals from bandpass filters 504 and506 are then subtracted in comparator circuit 508 to create adifferential signal for the microprocessor. A suitably configuredmicroprocessor then uses this differential signal to make suitableadjustments to the ultrasound-producing device.

[0107] In another embodiment of the present invention, a method forcontrolled enhancement of skin permeability by coupling fluid monitoringis disclosed. When a skin permeabilizing device, such as anultrasound-producing device, is applied to the skin, it is appliedthrough some coupling fluid, which may be a liquid, gel, or solid, tofacilitate transfer of the energy in the high frequency sound waves tothe skin. As the skin becomes more permeable, suitably-sized moleculesand ions in the coupling fluid begin to pass into and out of the skin.The method according to this embodiment takes advantage of the enhancedskin permeability that is the desired end point of this invention inorder to control the skin permeabilizing device. This method will beexplained in conjunction with the flow chart of FIG. 6.

[0108] In step 602 an initial concentration of a known substance isdetermined for the coupling medium. In practice, the coupling medium mayhave a known initial concentration of a known substance. That is, step602 will not require any additional measuring. The known substance canbe any substance (molecular or ionic) as long as its concentration inthe coupling medium is known. If, however, the substance is going to bepassed into the body, the substance should be one that is not harmful tothe body. The term benign is used herein to describe such a substance.Glucose and calcium are examples of substances that may be used in thisembodiment.

[0109] In step 604, the skin permeabilizing device is applied to thepatch of skin. In one embodiment, an ultrasound-producing device is usedas the skin permeabilizing device. Although the exact parameters ofultrasound are not the subject of this invention, according to oneembodiment, ultrasound having a frequency of about 20 kHz, and anintensity of about 10 W/cm² is used to enhance the permeability of thepatch of skin to be used for transdermal transport.

[0110] After the skin permeabilizing device has been turned on, in step606 the permeability of the patch of skin is monitored. According tothis embodiment, permeability monitoring is accomplished by monitoringchanges in the concentration of the known substance in the couplingmedium. That is, as the area of skin is subjected to the skinpermeabilizing device it will become permeable. As the area of skinbecomes permeable, molecules and ions begin to pass into the couplingmedium from inside the body and from the coupling medium into the bodydepending on the concentration gradient of the substance between thebody and the coupling medium. This concentration monitoring may be donein real time using an on-line sensor specifically programmed to detectand measure the concentration of the known substance.

[0111] In one embodiment, glucose is used as the known substance. Theconcentration of glucose is usually greater inside the body than in thecoupling medium unless the concentration in the coupling medium isartificially increased. Thus, when the skin becomes permeable, glucosemolecules will begin to pass into the coupling medium. In step 606,changes in the concentration of glucose in the coupling medium aremonitored to determine when the skin becomes permeable.

[0112] In another embodiment, mannitol is used as the known substance.Mannitol is a benign substance as that term is used in the context ofthis application. The concentration of mannitol in the coupling mediumis adjusted so that it is greater than the concentration of mannitol inthe body. When the skin becomes permeable, mannitol molecules with beginto pass from the coupling medium into the body, decreasing theconcentration of mannitol in the coupling medium. In step 606, thedecrease in the concentration of mannitol in the coupling medium ismonitored to determine when the skin becomes permeable.

[0113] In step 608, the skin permeabilizing device is controlled basedon the concentration measurements made in step 606. In one embodiment,the concentration measurements from the chemical analyzer are fed backto a microcontroller that is used to control the skin permeabilizingdevice. According to another embodiment, when the concentration of thesubstance being monitored reaches a predetermined value, the skinpermeabilizing device is turned off. If the concentration of thesubstance being monitored has not reached the predetermined value, themeasurement is repeated until the predetermined value is reached.

[0114] The predetermined value depends upon a number of factorsincluding, inter alia, the skin characteristics of the individual, theknown substance, and the frequency of the excitation source. As isapparent to one of ordinary skill in the art, a specific correlationbetween the change in concentration of the known substance being usedand skin permeability can be determined by conducting experiments andusing experimental data. The predetermined value is then determined on asubject-by-subject basis taking into account all appropriate factors aswell as any empirical data.

[0115] According to another embodiment, the intensity of the skinpermeabilizing device may be gradually scaled back as the point ofmaximum permeability enhancement is approached. In one embodiment, wherean ultrasound-producing device is used, as the concentration of thesubstance being monitored approaches 50% of the predetermined value,either the intensity or the duty cycle of the ultrasound may be reducedby a predetermined amount, such as 50%. This is done so that thepredetermined value is not “overshot” thereby increasing the risk ofskin damage. Additional controls are possible. For example, in anotherembodiment, the intensity may be scaled back when the concentration ofthe substance being monitored reaches 25%, 50% and 75% of thepredetermined value.

[0116] The rate of change in the concentration of the substance may alsobe used to determine a point at which the skin permeabilizing device isscaled back or discontinued. As the rate of change in the concentrationreaches a predetermined value, the intensity of the skin permeabilizingdevice may be gradually scaled back or discontinued, in a manner similarto that discussed above.

[0117] In another embodiment, skin permeability can be monitored bydetecting an electrical parameter of the coupling fluid. Morespecifically, as skin permeability increases, ions may pass into and outof the coupling medium. As ion concentration in the coupling mediumincreases or decreases, the electrical characteristics of the couplingmedium change. Therefore, the electrical characteristics of the couplingmedium can be used to monitor skin permeability using a methodology thatis a hybrid of that shown in FIGS. 2 and 6, and is set forth in FIG. 7.

[0118] In an initial step 702, a reference value for an electricalparameter is determined for the coupling medium. In practice, becausethe coupling medium has a known ionic composition, its electricalparameters should be known. In one embodiment, the coupling fluid has aknown concentration of calcium ions. Thus, this step should not requirean actual measurement. In another embodiment, the electrical parameterdetermined is conductivity, and in step 702, the conductivity of thecoupling medium is determined.

[0119] After the reference value for the electrical parameter isdetermined, in step 704 the skin permeabilizing device is turned on.Then in step 706 skin permeability is determined by monitoring changesin the electrical parameter of the coupling medium. This monitoring maybe accomplished using a simple meter. As the skin becomes permeable,depending upon the composition of the coupling medium, ions will passinto or out of the coupling medium and either increase or decrease theelectrical parameter of the coupling medium. In one embodiment, thecoupling medium has a known concentration of calcium ions that is lowerthan the concentration of calcium ions in the body. Therefore, as theskin becomes more permeable, calcium ions begin to pass from the bodyinto the coupling medium.

[0120] In step 708, the skin permeabilizing device is controlled basedon the monitoring measurements. In one embodiment, the monitoringmeasurements are fed back to a microcontroller that is used to controlthe skin permeabilizing device. In one embodiment, when the electricalparameter being monitored reaches is predetermined value, the skinpermeabilizing device is turned off. If the parameter being monitoredhas not reached the predetermined value, the measurement is repeateduntil the predetermined value is reached.

[0121] The rate of change in the parameter being monitored may also beused to determine a point at which the skin permeabilizing device isscaled back or discontinued. As the rate of change reaches apredetermined value, the intensity of the skin permeabilizing device maybe gradually scaled back or discontinued, in a manner similar to thatdiscussed above.

[0122] The predetermined value depends upon a number of factorsincluding, inter alia, the composition of the coupling medium, thesurface area of the patch of skin to which the skin permeabilizingdevice is applied, and the concentration of the particular ion beingused in the body. The predetermined value is determined on asubject-by-subject basis taking into account all appropriate factors andthe empirical data.

[0123] According to another embodiment, the intensity of the skinpermeabilizing device may be gradually scaled back as the point ofmaximum permeability enhancement is approached. In one embodiment, whereultrasound is used, as the parameter being monitored reaches 50% of thepredetermined value, either the intensity or the duty cycle is reducedby a predetermined amount, such as 50%. This is done so that thepredetermined value is not “overshot” thereby increasing the risk ofskin damage. Additional controls are possible. For example, in anotherembodiment, the intensity may be scaled back when the parameter beingmonitored reaches 25%, 50% and 75% of the predetermined value.

[0124] According to another embodiment of the present invention, anapparatus and method for regulating the degree of skin permeabilizationthrough a feedback system is provided. This apparatus and method may besimilar to what has been described above, with the addition of furtherregulation of the degree of skin permeabilization. In this embodiment,however, the application of the skin permeabilizing device is terminatedwhen desired values of parameters describing skin conductance areachieved. As the discussion proceeds with regard to FIG. 8, it should benoted that the descriptions above may be relevant to this description.

[0125] Referring to FIG. 8, a flowchart of the method is provided. Instep 802, a first, or source, electrode is coupled in electrical contactwith a first area of skin where permeabilization is required. Asdiscussed above, the source electrode does not have to make directcontact with the skin. Rather, it may be electrically coupled to theskin through the medium that is being used to transmit ultrasound. Inone embodiment, where an ultrasound-producing device is used as the skinpermeabilizing device, the ultrasonic transducer and horn that will beused to apply the ultrasound doubles as the source electrode throughwhich electrical parameters of the first area of skin may be measuredand is coupled to the skin through a saline solution used as anultrasound medium. In another embodiment, a separate electrode isaffixed to the first area of skin and is used as the source electrode.In still another embodiment, the housing of the device used to applyultrasound to the first area of skin is used as the source electrode.The source electrode can be made of any suitable conducting materialincluding, for example, metals and conducting polymers.

[0126] Next, in step 804, a second, or counter, electrode is coupled inelectrical contact with a second area of skin at another chosenlocation. This second area of skin can be adjacent to the first area ofskin, or it can be distant from the first area of skin. The counterelectrode can be made of any suitable conducting material including, forexample, metals and conducting polymers.

[0127] In order to get an accurate electrical reading, the counterelectrode should make sufficient contact with the skin. This can beachieved in a number of ways. In one embodiment, the counter electrodeis applied directly to the epidermis of the skin. That is, the counterelectrode is applied to an area of skin from which the stratum corneumhas been removed. The stratum corneum may be removed in a number ofways. According to one embodiment, the stratum corneum is removed bytape stripping. In another embodiment, sufficient electrical contactbetween the skin and the counter electrode is created by using a counterelectrode having a large surface area. More specifically, a conductivepolymeric path or metallic foil patch having an area much larger thanthe skin area exposed to the skin permeabilizing device is used. Thelarge area of the counter electrode in this embodiment decreases itsimpedances and allows accurate measurements of the electrical parameterof the area of skin exposed to the skin permeabilizing device. In onespecific embodiment a conductive band is wrapped around the subject'sarm and used as the counter electrode. In another embodiment, thecounter electrode may be placed in a handle of the skin permeabilizingdevice, to which a subject grasps during operation.

[0128] In another embodiment, the counter electrode surrounds the skinpermeabilizing device.

[0129] When the two electrodes are properly positioned, in step 806, aninitial conductivity between the two electrodes is measured. This may beaccomplished by applying an electrical signal to the patch of skinthrough the electrodes. In one embodiment, the electrical signalsupplied may have sufficient intensity so that the electrical parameterof the skin can be measured, but have a suitably low intensity so thatthe electrical signal does not cause permanent damage to the skin, orany significant electrophoresis effect for the substance beingdelivered. In one embodiment, a 10 Hz AC source is used to create avoltage differential between the source electrode and the counterelectrode. The voltage supplied should not exceed 500 mV, and preferablynot exceed 100 mV, or there will be a risk of damaging the skin. Inanother embodiment, an AC current source is used. The current source mayalso be suitably limited. The initial conductivity measurement is madeafter the source has been applied using appropriate circuitry. In oneembodiment a resistive sensor is used to measure the impedance of thepatch of skin at 10 Hz. In another embodiment, a 1 kHz source is used.Sources of other frequencies are also possible.

[0130] In step 808, a skin permeabilizing device is applied to the skinat the first site. Any suitable device that increases the permeabilityof the skin may be used. In one embodiment, ultrasound is applied to theskin at the first site. According to one embodiment, ultrasound having afrequency of 20 kHz and an intensity of about 10 W/cm² is used toenhance the permeability of the patch of skin to be used for transdermaltransport.

[0131] In step 810, the conductivity between the two sites is measured.The conductivity may be measured periodically, or it may be measuredcontinuously. The monitoring measurements are made using the sameelectrode set up that was used to make the initial conductivitymeasurement.

[0132] In step 812, mathematical analysis and/or signal processing maybe performed on the time-variance of skin conductance data. Experimentswere performed on human volunteers according to the procedure above,with ultrasound used as the method of permeabilization. Ultrasound wasapplied until the subjects reported pain. Skin conductivity was measuredonce every second during ultrasound exposure. After plotting theconductance data, the graph resembled a sigmoidal curve. The conductancedata was in a general sigmoidal curve equation:$C = {{Ci} + \frac{\left( {C_{f} - C_{i}} \right)}{1 + ^{S{({t - t^{*}})}}}}$

[0133] where:

[0134] C is current;

[0135] C_(i) is current at t=0;

[0136] C_(f) is the final current;

[0137] S is a sensitivity constant;

[0138] t* is the exposure time required to achieve an inflection point;and

[0139] t is the time of exposure.

[0140]FIG. 9 shows the time variation of the skin conductance whilebeing exposed to ultrasound. The curve is a sigmoidal curve and can befitted to the above equation. The line shown in FIG. 9 corresponds to afit to the above equation. The values of fitted parameters were obtainedand are plotted. The value of t* corresponds to an exposure timerequired to achieve an inflection point (a point where the slope of thecurve shown changes sign). The inflection time approximately indicatesthe time required to achieve half the total exposure.

[0141]FIG. 10 shows a relationship between the inflection time and thepain time on various volunteers. The data shows that the time to pain isproportional to the time to the inflection point on human volunteers. Inthis figure, R2 is the correlation coefficient, where a R2=1 indicates100% correlation of the experimental data to the predicted values. Basedon this data, a method can be developed to predict the requiredultrasound exposure time.

[0142] Referring to FIGS. 11 and 12, a flowchart depicting a method ofdetermining when to terminate the application of ultrasound, andcorresponding example graphs, are provided. In step 1102, A/D conversionis performed on the conductivity data. This results in a graph similarto the one in FIG. 12a. Next, in step 1104, filtering is performed onthe digital data. As shown in FIG. 12b, the filtered data has a smoothercurve than the unfiltered data of FIG. 12a. Next, in step 1106, theslope of the curve is calculated. In step 1108, the maximum value forthe slope is saved. If the current value for the slope is greater thanthe maximum value that is saved, the maximum value is replaced with thecurrent value. Next, in step 1110, if the slope is not less than orequal to the maximum value, the process returns to step 1102 to wait fora peak. If the slope is less than or equal to the maximum value, in step1112 the process detects a peak, or point of inflection, shown in FIG.12c, then, in step 1114, terminates the application of ultrasound to theskin.

[0143] In one embodiment, the detection of the peak may be validated.This may be provided to ensure that the “peak” detected, in step 1112,was not noise, but was actually a peak.

[0144] In other embodiments, ultrasound may be applied even after theinflection point is reached. In one embodiment, ultrasound is appliedfor a predetermined time. This predetermined time may be based on apercentage of the time to reach the inflection point. For example, oncethe inflection point is reached, ultrasound continues to be applied foran additional 50% of the time it took to reach the inflection point.Thus, if it took 14 seconds to reach the inflection point, ultrasound isapplied for an additional 7 seconds. Other percentages may be used, andthis percentage may be based on factors including pain threshold andskin characteristics.

[0145] In another embodiment, ultrasound is applied until the slopedecreases to a certain value. Referring again to FIG. 11, after theinflection point is reached, the slope decreases as ultrasound isapplied. Thus, ultrasound may be applied until the slope decreases by apercentage, such as 50%, or to a predetermined value. As above, thisdetermination is flexible and may vary from individual to individual.

[0146] In another embodiment, the current at the inflection point ismeasured, and then a percentage of this current is still applied. Forexample, if the inflection point is reached at 40 μamps, an additional10% of this, for a total of 44 μamps, may be reached. Again, thisdetermination is flexible and may vary from person to person.

[0147] Referring again to FIG. 8, in step 814, the parameters describingthe kinetics of skin conductance changes are calculated. Theseparameters include, inter alia, skin impedance, the variation of skinimpedance with time, final skin impedance, skin impedance at inflectiontime, final current, exposure time to achieve the inflection time, etc.

[0148] In step 816, the skin permeabilizing device applied in step 808is terminated when desired values of the parameters describing skinconductance are achieved.

EXAMPLE

[0149] In vitro experiments were performed in accordance with a methodaccording to one embodiment of the present invention. Pig skin wasmounted on a diffusion cell and was exposed to ultrasound using 1%Sodium Lauryl Sulfate in water as a coupling medium. Skin conductancewas measured by placing two electrodes across the skin. The impedanceswere measured at two frequencies: at 10 Hz, which is near the ultrasoundrange, and 1 kHz. The impedances measured at the frequencies differed byabout 25 fold when the skin was impermeable. Upon sonication, thedifference between the impedances at the two frequencies decreased. Thedifferential impedance between the two frequencies decreased with time.When the skin was highly permeable, the impedances at two frequenciesdiffered by only about 20%. SLS was removed from the chamber and thechamber was dried. A gel was placed in the chamber in contact with theskin. The gel was prepared by mixing glucose reagent from the Sigma™ kit315 (10% by weight) into polyvinyl alcohol solution (20% by weight) inPBS. The gel was kept in the freezer to allow cross linking. The gel wasclear in the beginning, and changed color to red when it came in contactwith glucose.

[0150] 2. Extraction and Analysis of At Least One Analyte in Body Fluid

[0151] According to another embodiment of the present invention,ultrasound may be used to extract body fluids through or out of skinthat has its permeability increased. Referring to FIG. 13, a flowchartdepicting a method for extraction and analysis of at least one analytein a body fluid according to one embodiment of the present invention isdisclosed. In step 1302, the permeability of the skin is increased. Thismay be accomplished by any suitable method for increasing thepermeability of the skin, such as iontophoresis. In one embodiment, thepermeability of the skin may be increased through the application ofultrasound.

[0152] As used herein, the term “interstitial fluid” may include lymph,interstitial fluid, and serum that may be extracted from the body. It isalso used to describe components of interstitial fluid.

[0153] In step 1304, interstitial fluid is extracted transdermally fromthe surface of the skin. Extraction can be performed after sonication orother permeation methods using a wide variety of different forces. Theseforces may include physical forces, chemical forces, biological forces,vacuum pressure, electrical, osmotic, diffusion, electromagneticultrasound, cavitation, mechanical, thermal, capillary forces, fluidcirculation across the skin, electro-acoustic, magnetic,magneto-hydrodynamic, acoustic, convective dispersion, photo acoustic,by rinsing body fluid off skin, or by any combination of these forces.

[0154] Spatial and/or temporal positive and/or negative pressuremodulation may be used. In spatial modulation, positive pressure isapplied to an area of the skin, while a vacuum is applied to anotherarea, assisting in the extraction of body fluid. In temporal modulation,vacuum and positive pressure alternate at about the same area of skin,assisting in the extraction of body fluid. The application of eitherspatial or temporal modulation may be continuous or discontinuous, andthey may be applied separately or in combination.

[0155] In one embodiment, vacuum pressure may be applied to extract bodyfluid. Vacuum pressure may be applied continuously, or it may be applieddiscontinuously. When applied discontinuously, the vacuum may be appliedin a pulsed fashion. A material that maintains the surface configurationof the skin (e.g., flat, convex, or concave), such as mesh, membrane,perforated metal, or other porous material, may be applied between thevacuum pressure and the skin while the vacuum pressure is applied. Thevacuum can act through these structures and can be generatedmechanically, electro-mechanically, chemically, or electrochemically. Inanother embodiment, the vacuum can be applied in such a manner so as tomaintain the skin surface configuration with the vacuum alone.

[0156] In another embodiment, a chamber that is applied to skin can havea design (configuration and material properties) to localize highpressure gradient across skin and/or other tissues.

[0157] In another embodiment electrical, forces may be applied.Electrical forces may be iontophoretic, electro-osmotic, or may beelectroporation. A gel with an electric charge also may be applied, inorder to encourage the absorption and evacuation of body fluid andcomponents thereof.

[0158] In another embodiment, osmotic forces may be used. A gel orsolution may be applied to the skin surface in order to encourageosmosis.

[0159] In another embodiment, ultrasound may be used to pump body fluidand fluid components, to levitate, to activate gas bodies, to producecyclic impulse mechanical stress to the skin, to create microstreaming,to increase temperature, or to set up standing waves. Single or multiplesources of ultrasound may be used in combination with variouscharacteristics of ultrasound, e.g., different frequencies, intensities,or coupling media, in order to encourage the extraction of body fluid.

[0160] In another embodiment, mechanical forces may used to extract bodyfluid. These forces may be achieved by, inter alia, a roller, asqueezer, a stretcher, iris compressor/tensioner device, etc. toincrease the volume of the body fluid that is extracted.

[0161] In one embodiment, a tensioner is used to extract body fluid.Referring to FIG. 14, which depicts an embodiment of a tensioner,tensioner 1402 consists of a convex geometry held against the skin 1404.By pressing tensioner 1402 against skin 1404, body fluid may becollected within cavity 1406 of tensioner 1402.

[0162] In another embodiment, thermal forces may be used to extract bodyfluid. The skin temperature may be increased using electricity,chemical, ultrasonic, or optical energy sources or methods and/orutilize temperature sensitive polymers to swell or contract a gel,membrane, and/or solid to encourage the absorption and evacuation ofbody fluid and components thereof. Temperature sensitive polymers may beused to move a piston or membrane to push or suck fluid. Examples ofsuch polymers include, inter alia, poloxymers.

[0163] In another embodiment, chemical forces are used Chemicalsubstances may be used to augment convective and/or diffusive forces asa means to extract additional body fluid, and/or to enhance transportand/or accumulation of body fluids at specific body sites. A hydrogelwith an incorporated trapped, or immobilized bioactive molecules such asenzyme would allow for extraction by osmosis into a sensing scaffold.

[0164] In another embodiment, pH/ionic forces may be used These forcesmay be used to change the material properties and characteristics, e.g.,hydrophilic material to a hydrophobic material. A pH/ionic sensitivemembrane and/or gel may be swollen and contracted in order to encouragethe absorption and evacuation of body fluid and components thereof.

[0165] In another embodiment, capillary forces may be used. These forcesmay be used to assist in fluid transport across skin pores.

[0166] Referring again to FIG. 13, in step 1306, the body fluid andcomponents thereof are collected. This collection may be accomplished byabsorption, adsorption, phase separation, mechanical forces, electricalforces, chemically induced forces, or a combination thereof. Preferably,a humid environment is created and maintained in order to controlevaporation of analytes during extraction. The collected volume of bodyfluid may be the same as the volume extracted, or it may be a fixedconstant volume.

[0167] In one embodiment, absorption or adsorption may be used. In thisembodiment, the body fluid may be collected into a gel, which contains acaptive enzyme. A polymeric, metallic, or ceramic screen, scaffold,mesh, or membrane, or a combination may be used to do this. Thesematerials may also be a component of a sensor.

[0168] In one embodiment, phase separation may be used. Body fluid maybe isolated by combining the fluid with an appropriate densityimmiscible fluid. The body fluid may be collected into a conicalchamber.

[0169] Another use of phase separation is achieved by first applying ahydrophobic coating on the skin prior to the extraction step. After theextraction, body fluid is present in the form of droplets on thehydrophobic coating.

[0170] In another embodiment, mechanical forces may be used to collectbody fluids. This includes forces such as vacuum, pressure, and acousticforces. Dispersed body fluid may be collected over a greater area to asmaller area using a microfluidic channel against the skin. A means toevacuate the fluidic path may include the introduction of a liquidand/or gas. This means to evacuate may be applied to all collectionprocesses, and not just mechanical collection.

[0171] In another embodiment, electrical collection may be used. In thisembodiment, solid, liquid droplets, or gas are charged and transported(moved) from skin to a sensor or to a collecting compartment usingelectrical forces.

[0172] In another embodiment, chemical collection may be used. Ahydrophilic gel may be used to collect body fluids. The materialproperties and characteristics may be changed, e.g., hydrophilicmaterial to a hydrophobic material, in order to encourage the absorptionand evacuation of body fluid and components thereof.

[0173] In another embodiment, capillary collection may be used. Bodyfluid may be collected into a capillary or capillaries. This allows forquantitative volume or a method to move fluid to a sensor. The capillaryor capillaries may be filled with multiple fibers to increase thesurface area on which a liquid's adhesive forces can act. This methodmay be used in conjunction with a chemical substance and/or otherdriving forces.

[0174] In step 1308, the concentration of analytes in body fluid issensed. Sensing the concentration of an analyte present in body fluidmay be accomplished by employing electrochemical, optical, acoustical,biological, and enzymatic technology in combination or alone. A sensoror sensors can be disposable, replenishable, discrete, or continuous.

[0175] According to one embodiment, a sensing device may have a sensoror sensors capable of detecting more than one analyte. If one or more,or a combination of several analytes exists in stable and/or predictablephysiologic concentrations, the ratio of one analyte to the other wouldallow for concentration detection and self-calibration. Neither thevolume of the body fluid or the dilute volume needs be known.

[0176] A sensing device presented with a known volume of body fluid(undiluted) and a known volume of diluent would not require frequentcalibration.

[0177] In one embodiment, body fluid is extracted and optical analysisis performed on the body fluid.

[0178] In another embodiment, the body fluid is extracted andelectrochemical analysis is performed on the body fluid.

[0179] In another embodiment, the body fluid is extracted and theacoustical emission of an analyte undergoing a chemical reaction isdetected and analyzed.

[0180] In another embodiment, the body fluid is extracted, and livingcells may be used to sense a concentration of an analyte in body fluid.

[0181] In another embodiment, the body fluid is extracted, and thermalanalysis is performed on the body fluid.

[0182] In step 1310, information is provided for the user interface. Auser interface may provide features for both daytime and nighttimemonitoring. In one embodiment, this may include alarms for high/lowanalyte concentrations, may provide access to trends and history, andmay enable a prediction of future concentration values. The userinterface may provide the ability to download history. Other conveniencefeatures, such as a low battery indicator, may be included in the userinterface. The battery may be solar, nickel cadmium, standard alkaline,or lithium ion.

[0183] In another embodiment, after the permeability of the skin isincreased, the presence of the analyte may be sensed without extraction.Infrared light, for example, may be used to sense the presence of ananalyte, with less interference from H₂0.

[0184] In another embodiment, after the permeability of the skin isincreased, at least one analyte is permitted to passively diffusethrough the skin. The analyte may be collected in a gel used as acollecting device, and a sensing device, attached to the gel, may beused to sense the presence of the analyte.

[0185] In another embodiment, additional methods for generatingcavitation and convectional flow may be applied with, before, after, orinstead of the ultrasound application for skin permeabilization and/orextraction and/or collection steps. These methods include the use of apropeller, fly wheel, transverse needle, and local shear inducedpermeabilization.

[0186] In another embodiment, the non-invasive method disclosed hereinmay be used to determine the level of blood glucose. Referring to FIG.15, in step 1502, the permeability of the skin is increased. This may beachieved by any suitable method. Preferably, ultrasound is applied asdiscussed above.

[0187] In step 1504, the interstitial fluid is extracted from the skin.This may be accomplished by any suitable method, including thosediscussed above. Preferably, a vacuum is applied to extract theinterstitial fluid from the skin.

[0188] In step 1506, the interstitial fluid is collected. This may beaccomplished by any suitable method, including those discussed above.Preferably, the interstitial fluid is collected into a gel containingglucose sensitive reagents. The gel may change color when it comes incontact with glucose.

[0189] In step 1508, the color change of the gel is monitored todetermine the glucose concentration in the interstitial fluid.

[0190] In another embodiment, ultrasound may be used for detection(evaluation, follow up treatments) of skin and/or other subcutaneousabnormalities presented by pathological concentrations of specificanalytes, or detection of specific components administered to the siteand detection of their elimination or conversion (psoriasis, skinmalignancies, etc.). This approach may be used, for example, inextracting analytes or reagents from skin-affected sites (lesionplaques), tumors, etc.

[0191] In another embodiment, the delivery and/or removal of endogenousand non-endogenous components from the skin by the application by aforce is disclosed. Forces, such as ultrasound, electrical, magnetic,capillary, mechanical, chemical, electromagnetic, osmotic, concentrationgradient, or combinations thereof may be used in applications such as,inter alia, removal of residual surfactant, cavitation enhancers, tattoobleach, and botox (remove from forehead, and neck lines, reducesweating).

[0192] In another embodiment, sensing components may be delivered intothe skin to analyze interstitial fluid components in situ. The sensingcomponents may also be delivered into the skin to measure emittedproducts or reagents of any sensing reaction (chemical or enzymatic).

[0193] 3. Sonophoretic Drug Delivery

[0194] A drug is defined as a therapeutic, prophylactic, or diagnosticmolecule or agent, that may be in a form dissolved or suspended in aliquid, solid, or encapsulated and/or distributed in or within micro ornanoparticles, emulsion, liposomes, or lipid vesicles. Drug delivery isdefined as the delivery of a drug into blood, lymph, interstitial fluid,cells, tissues, and/or organs, or any combination thereof.

[0195] Referring to FIG. 16, an active patch drug delivery apparatus1602 that is attached to skin 1600 is depicted. Drug delivery apparatus1602 includes patch 1604. Patch 1604 includes adhesive 1610, drugmolecules 1612 and transducer 1614. Patch 1604 is an active patch.Adhesive 1610 acts as an attaching device. Alternatively, the attachingdevice may be a vacuum, band, or strap. As transducer 1614 oscillates,the permeability of skin 1600 is increased in accordance with thepresent invention and drug molecules 1612 are delivered to and/orthrough skin 1600, or/and after skin 1600 is permeabilized, drugmolecules 1612 are transported through skin 1600 to the capillaries andblood vessels below skin 1600. A limiting step membrane 1613 may belocated between skin 1600 and drug molecules 1612.

[0196] Transducer 1614 preferably operates at a frequency in the rangeof between 20 kHz to 2.5 MHz, using appropriate electrical signalgenerators and amplifiers. Transducer 1614, more preferably, isoperating at a frequency in the range of between 20 and 200 kHz. Otherultrasound parameters include, but are not limited to, amplitude, dutycycle, distance from the skin, coupling agent composition, andapplication time and may be varied to achieve sufficient enhancement oftransdermal transport. The intensity preferably varies from 0 to 20W/cm². Further, transducer 1614 may be configured as a cylinder, ahollow cylinder, a hemispherical configuration, conical configuration,planer configuration or rectangle configuration. Transducer 1614 mayalso consist of an array of acoustic elements that are swept in time.Transducer 1614 may be comprised of quartz, PVDF, ceramic including PZTand screen printed ceramic, magnetostrictive, or composite materialincluding molded ceramic and benders. Transducer 1614 may be used alone,or in conjunction with other forces, or contributors, to enhance drugdelivery. These other forces, or contributors, include, but are notlimited to, a magnetic field including electromagnetic forces, anelectrical current or iontophoresis, mechanical skin manipulation,chemical enhancement, heat, and osmotic forces.

[0197] Transducer 1614 administers ultrasound preferably at frequenciesof less than or equal to about 2.5 MHz, preferably at a frequency thatis less than 1 MHz, and more typically in the range of about 20 to 100kHz. Exposures to ultrasounds from transducer 1614 are typically betweenabout 5 seconds and about 10 minutes continuously, but may be shorterand/or pulsed, for example, at 100 to 500 msec pulses every seconds fora time sufficient to permeabilize the skin. The ultrasound intensity isof a level that preferably does not raise skin 1600's temperature morethan about 1 to 2 degrees Centigrade and does not cause permanent damageto the skin. The intensity typically is less than 20 W/cm², preferablyless than 10 W/cm². Intensity in time of application is inverselyproportional to exposure time, so that high intensities are applied forshorter period of times in order to avoid skin damage. It should benoted that although normal low range ultrasound is 20 kHz, comparableresults are achieved by varying the frequency to less than 20 kHz, orinto the sound region.

[0198] The time needed for permeabilization is dependant upon thefrequency and intensity of the ultrasound from transducer 1614 and thecondition of skin 1600. At 20 kHz, for example, an intensity of 10W/cm², with a duty cycle of 50 percent, skin 1600 is permeabilizedsufficiently in about 5 minutes if skin 1600 is on a human forearm.

[0199] Permeabilizing ultrasound may be applied for a predeterminedamount of time or may be applied only until permeabilization isattained. Because skin 1600 characteristics or properties may changeover time, based on aging, diet, stress, and other factors, it may bepreferable to measure permeability as ultrasound is applied to minimizethe risk of skin 1600 damage. Several methods may be used to determinewhen sufficient permeabilization has been reached. One method measuresrelative skin conductivity at the permeabilization site versus areference point. These measurements are performed by applying a small ACor DC electric potential across two electrically isolated electrodes incontact with skin 1600. Electric current flowing through theseelectrodes is measured using an ammeter and skin 1600 resistance ismeasured using the values of the potential and current. Drug deliverypatch apparatus 1602 may serve as one of the electrically isolatedelectrodes in contact with skin 1600. Preferably, drug delivery patchapparatus 1602 permeabilizes skin 1600 prior to the conductivity tests.

[0200] Another way to determine when sufficient permeabilization hasbeen reached is to measure conductivity. Fully permeabilized skin has aresistance of no more than about 5 kΩ measured across approximately 1.7cm². Another method is to detect and/or quantitate the transdermalmovement of an analyte, such as creatinine, calcium or total ions, thatis present in interstitial fluid in a fairly constant amount, and may beused either to calibrate the concentration of analyte to be extractedand quantified, or as a measure of permeabilization. The higher theconstant analyte flux, the greater degree of permeabilization. Thedegree of permeability also may be monitored using a sensor attached todrug delivery patch apparatus 1602 that determines the concentration ofdrug molecules 1612 being delivered or an analyte being extracted. Asthe permeability increases, the drug concentration within drug deliverypatch 1602 decreases.

[0201] Drug delivery patch apparatus 1602 also may be applied topretreated skin 1600. In other words, permeabilization of skin 1600 isalready achieved. Drug delivery patch apparatus 1602 is placed overpretreated skin 1600 to deliver drug molecules 1612. Any known devicemay bemused to pre-treat skin 1600, including, but not limited to,physical forces, chemical forces, biological forces, vacuum pressure,electrical forces, osmotic forces, diffusion forces, electromagneticforces, ultrasound forces, cavitation forces, mechanical forces, thermalforces, capillary forces, fluid circulation across the skin,electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces,acoustic forces, convective dispersion, photo-acoustic forces, byrinsing body fluid off skin, and any combination thereof.

[0202] Drug molecules 1612 include a variety of bio-active agents,including protein and peptides. Other materials include nucleic acidmolecules such as vaccines including therapeutic proteins, syntheticorganic and inorganic molecules including anti-inflammatories,anti-virals, anti-fungal, antibiotics, and local anesthetics, andsaccharides and polysaccharides. Drug molecules 1612 may be administeredin an appropriated pharmaceutically acceptable carrier having anabsorption coefficient, similar to water, such as an aqueous gels,ointment, lotion, or suspension. Drug molecules 1612 also may becontained with adhesive 1610 that attaches to skin 1600. Further, drugmolecules 1612 also may be encapsulated or suspended in a liquid, gel,or solid matrix within patch 1604.

[0203] Drug delivery patch apparatus 1602 also includes a battery 1616.Battery 1616 acts as a power source for transducer 1614. Battery 1616provides a relatively high energy burst. Drug delivery patch apparatus1602 also includes electronic coupling 1618 that serves as the driveelectronics for drug delivery patch apparatus 1602. Drug delivery patchapparatus 1602 also includes user interface 1620.

[0204] In one embodiment, patch 1604 includes transducer 1614, drugmolecules 1610, and adhesive 1610. In another embodiment, patch 1604includes transducer 1614, drug molecules 1612, adhesive 1610, battery1616, electronic coupling 1618, and user interface 1620. In anotherembodiment, patch 1604 includes transducer 1614, drug molecules 1612,adhesive 1610, and battery 1616. In another embodiment, adhesive 1610 isto the side of transducer 1614 and drug molecules 1612.

[0205] Battery 1616, electronic coupling 1618, and user interface 1620,may be located elsewhere on a user and in communication with patch 1604via hard wire or telemetry In another embodiment, user interface 1620may be located elsewhere on the user and is in communication with patch1604 via hard wire, telemetry, infra-red, or fiber optic means. Thus,the elements of drug delivery apparatus 1602 may be detachable andportable from each other. Further, any of the components of drugdelivery apparatus 1602 may be disposable or reusable. For example,patch 1604, which includes transducer 1614, drug molecules 1612 andadhesive 1610 may be disposed after detachment from skin 1600. However,battery 1616, electronic coupling 1618, and user interface 1620 may bere-usable with further patches 1604.

[0206] In one embodiment, transducer 1614 operates alone to push drugmolecules 1612 through and to skin 1600. Alternatively, drug deliverypatch apparatus 1602 and transducer 1614 may operate in conjunction witha driving force that further facilitates the transdermal transport ofdrug molecules 1612. These forces include, but are not limited tophysical forces, chemical forces, biological forces, vacuum pressure,electrical forces, osmotic forces, diffusion forces, electromagneticforces, ultrasound forces, cavitation forces, mechanical forces, thermalforces, capillary forces, fluid circulation across the skin,electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces,acoustic forces, convective dispersion, photo-acoustic forces, byrinsing body fluid off skin, and any combination thereof.

[0207] Referring to FIG. 17, an embodiment of transducer 1614 isdepicted. Transducer 1614 may be an array of acoustic elements that areswept in time as ultrasound is applied to drug molecules 1612, andthrough adhesive 1610 to skin 1600. Acoustic elements 1700 comprisetransducer 1614. Elements 1700 are depicted as squares within a largersquare. Elements 1700 are not limited to this configuration and may beconfigured as a cylinder, a hollow cylinder, hemispherical, conical,planer, rectangular. Each acoustic element of elements of 1700 may beswept individually or within a group as transducer 1614 is activated.For example, element A activates, followed by elements B and E, thenfollowed by elements C, F, and I, and so on. Element P may be activatedlast as transducer 1614 is swept. Further, acoustic elements 1700 maycomprise fingers. Referring to FIG. 17, a finger may be depicted aselements A, E, I, and M. Each finger may be activated or swept in time.Acoustic elements 1700 may be configured to channel the ultrasoundenergy from transducer 1614 to a specified area in 100 smaller than thearea of transducer 1614.

[0208] Referring to FIG. 18, patch 1604 and user interface 1620 arecoupled to feedback mechanism 1802. Feedback mechanism 1802 may bedetachable from user interface 1620. Alternatively, feedback mechanism1802 may be contained within user interface 1620. Thus, feedbackmechanism 1802 may be contained within drug delivery patch apparatus1602. Feedback mechanism 1802 provides for programming of drug deliveryrates or pre-set doses of drug molecules 1612. Feedback mechanism 1802also may provide memory to record or display historical delivery data touser interface 1620. Feedback mechanism 1802 communicates the on time oftransducer 1614 to user interface 1620 for display to the user. Feedbackmechanism 1802 also may provide alarms for low drug molecules 1612and/or low power in battery 1616. Thus, feedback mechanism 1802 alerts auser via a user interface 1620 that drug molecules 1612 and patch 1604needs to be replenished or that drug delivery patch apparatus 1602 islow on power.

[0209] Feedback mechanism 1802 also may monitor the amount of drugmolecules 1612 delivered via transdermal transport. Feedback mechanism1802 also may monitor the amount of ultrasonic energy, or other drivingforces listed above, applied to skin 1600 by transducer 1614. Limits maybe set in feedback mechanism 1802 to limit the ultrasound energy fromtransducer 1614 so as to no irritate or damage skin 1600. Feedbackmechanism 1802 also may monitor the concentration of drug molecules 1612remaining in patch 1604. Feedback mechanism 1802 also may monitor theconcentration of drug molecules or analytes in the interstitial fluid,blood, and other body fluids. Feedback mechanism 1802 also may monitorthe amount of cavitation produced by the application of ultrasoundenergy. Feedback mechanism 1802 also may monitor the degree ofphysiological effects such as blood pressure, EMG, EEG, and ECT feedbackin order to measure delivery of drug molecules 1612. Feedback mechanism1820 also may provide connections with additional patches or testingdevices in order to perform conductivity tests.

[0210] 4. Transdermal Vaccination by Sonophoresis

[0211] Generally, vaccines are administered for the prevention,amelioration or treatment of infectious diseases. Vaccines are commonlyused to provide immunity from diseases such as influenza, poliomyelitis,varicella zoster (chicken pox), measles, as well as several otherdiseases.

[0212] A vaccine is generally made from an antigen isolated or producedfrom the disease-causing microorganism. An antigen is defined as“anything that can be bound by an antibody.” This can be an enormousrange of substances from simple chemicals, sugars, small peptides tocomplex protein complexes, such as viruses. The small antigens are not,however, immunogenic in themselves, and need to be coupled to a carrierto elicit an immune response.

[0213] Typically, the vaccine is delivered to the bloodstream by aninvasive method, such as an injection. The B cells in the blood streamrespond to the antigen by producing antibodies. These antibodies bind tothe antigen to “neutralize,” or inactivate it. Memory cells are alsoproduced, and remain ready to mount a quick protective immune responseagainst subsequent infection by the same disease-causing agent.

[0214] Immunization is the process of causing immunity by injectingantibodies or provoking the body to make its own antibodies against acertain microorganism. Immunization may be a result of a vaccination.

[0215] As discussed above, the use of ultrasound to facilitatetransdermal transport is known. The mechanism by which ultrasound isused to facilitate transdermal transport has differed. In the context oftransdermal delivery systems, ultrasound was initially a driving forcethat essentially pushed drugs through the skin and into the circulatorysystem. Ultrasound also increases the permeability of the skin. In otherwords, application of ultrasound having a particular frequencydisorganizes the lipid bi-layer in the skin, thereby increasing thepermeability of the skin. In this context, drugs may be delivered to thebody through the skin, or an analyte may be extracted from the bodythrough the skin. System and methods for the application of ultrasoundto enhance the permeability of skin, as well as the extraction of bodyfluids are discussed, above.

[0216] Although the permeability of the skin is increased by theapplication of ultrasound, a driving force is still required fortransdermal transport, but the required intensity of the driving forceis decreased. For example, a concentration gradient is generally asufficient driving force for transdermal transport through skin whosepermeability has been enhanced using ultrasound. The permeabilityenhancement that results from the application of ultrasound is due atleast in part, to cavitation that is caused by the ultrasound.

[0217]FIG. 19 depicts a method for transdermal vaccination bysonophoresis according to one embodiment of the present invention.Referring to FIG. 19, in step 1902, the permeability of the skin isincreased. This may be achieved by several methods, including thosediscussed above.

[0218] In one embodiment, ultrasound may be applied at about 10 W/cm²,with a duty cycle of about 50%. Ultrasound may be applied at a distancefrom the skin of about 0.5 mm to 1 cm, and for an application time offrom about 30 seconds to about 5 minutes.

[0219] A coupling medium may be used between the transducer and theskin, and may contain aqueous or non-aqueous chemicals including, butnot limited to, water, saline, alcohol, including ethanol andisopropanol (1-100% mixtures with saline), surfactants, fatty acids suchas linoleic acid (0.1-2% mixtures in ethanol-water (50:50) mixture),azone (0.1-10% mixtures in ethanol-water (50:50) mixture), 01-50%polyethylene glycol in saline, 1-100 mM EDTA, EGTA, or 1% SLS and silicaparticles. The coupling media provide effective transfer of ultrasoundenergy from transducer to the skin. Appropriate mixtures of thesecoupling media may also enhance cavitation activity inside, on thesurface, or near the skin, thus inducing more effective transport ofmolecules across the skin.

[0220] In step 1904, after the permeability of the skin is increased,sonication is terminated, and a vaccine is provided on the permeatedskin. In one embodiment, the vaccine may be incorporated into atransdermal patch. Other forms of the vaccine, such as gels and liquids,may also be used.

[0221] The vaccine may comprise as the active ingredient a peptide,protein, allergen, or other antigen, or DNA encoding any of theforegoing and may also include other adjuvants normally employed. Thesevaccines may be used as cancer vaccines, tetanus vaccines, etc.

[0222] In step 1906, the vaccine is delivered to the skin cells. In oneembodiment, the vaccine is delivered to skin cells, including Langerhanscells, dendritic cells, and keratinocytes.

[0223] In one embodiment, the vaccine is delivered to the Langerhanscells. The Langerhans cells are the cells responsible for capturing avaccine and presenting it to the Lymphatic system, and eliciting animmune response. The vaccine may be delivered to other cells to illicitan immune response.

[0224] In one embodiment, the vaccine may diffuse to the skin cells,including Langerhans cells, dendric cells, and keratinocytes. Once thevaccine is received by the skin cells, the vaccine is transported to thelymph nodes efficiently, increasing the efficiency of vaccination.

[0225] In another embodiment, the vaccine is transported transdermallythrough, in, or into the skin and into the bloodstream, wherein it actsas if it were injected in a conventional manner.

[0226] In another embodiment of the present invention, the vaccine isprovided simultaneously with the application of ultrasound. Theultrasound in this embodiment is used both to permeabilize the skin, aswell as and to deliver the vaccine transdermally to the Langerhanscells. The ultrasound acts as a driving force. Examples of usingultrasound to transport drugs from a patch are discussed above.

[0227] In another embodiment of the present invention, ultrasound isapplied to the skin to increase the permeability of the skin. Once thevaccine is provided, additional driving forces are provided to deliverthe vaccine to the body. Examples of driving forces include, inter alia,physical forces, chemical forces, biological forces, vacuum, electricalforces, osmotic forces, diffusion forces, electromagnetic forces,ultrasound forces, cavitation forces, mechanical forces, thermal forces,capillary forces, fluid circulation across the skin, electro-acousticforces, magnetic forces, magneto-hydrodynamic forces, acoustic forces,convective dispersion, photo acoustic forces, and any combinationthereof.

[0228] In another embodiment, ultrasound can be used to induceirritation and inflammation of the skin. Inducing irritation andinflammation may make the vaccine placed on the skin more effective ininducing an immune response.

[0229] In another embodiment, chemical enhancers may be used to increasethe permeability of the skin.

[0230] Although the present invention has been described in detail, itshould be understood that various changes, substitutions, andalterations can be made without departing from the intended scope asdefined by the appended claims.

What is claimed is:
 1. A method for enhancing transdermal transport, comprising: increasing a permeability of an area of a membrane with a permeabilizing device; monitoring the permeability of the area of membrane; transporting a substance into and through the area of membrane.
 2. The method of claim 1, wherein the step of increasing a permeability of an area of membrane with a permeabilizing device comprises: applying electricity to the area of membrane; measuring at least one electrical parameter of the area of membrane; and controlling the permeabilizing device based on the at least one electrical parameter.
 3. The method of claim 1, wherein the step of increasing a permeability of an area of membrane with permeabilizing device comprises: creating a volume of fluid adjacent the area of membrane, said fluid having an initial concentration of a first substance; and applying the permeabilizing device to the area of membrane.
 4. The method of claim 3, wherein the step of monitoring the permeability of the area of membrane comprises: monitoring changes in the concentration of the first substance; and controlling the permeabilizing device based on the changes in the concentration of the substance.
 5. The method of claim 1, wherein the step of increasing a permeability of an area of membrane with a permeabilizing device comprises: creating a volume of fluid adjacent the area of membrane; determining a reference value for an electrical parameter of the volume of fluid; and applying the permeabilizing device to the area of membrane.
 6. The method of claim 5, wherein the created volume fluid is selected from the group consisting of a liquid, a gel, and a solid.
 7. The method of claim 5, wherein the step of monitoring the permeability of the area of membrane comprises: monitoring changes in the electrical parameter of the volume of fluid; and controlling the permeabilizing device based on the changes in the electrical parameter of the volume of fluid.
 8. The method of claim 1, wherein the step of increasing a permeability of an area of membrane with a permeabilizing device comprises: providing a first electrode in electrical contact with a first area of membrane; providing a second electrode in electrical contact with a second area of membrane; measuring an initial conductivity between said electrodes; and applying the permeabilizing device to said first area of membrane.
 9. The method of claim 8, wherein the step of monitoring the permeability of the area of membrane comprises: measuring a second conductivity between said first and second electrodes; processing said initial conductivity and said second conductivity to establish information on a time-variation of membrane conductance; calculating at least one parameter describing a kinetics of membrane conductance changes responsive to said information; and terminating said application of said permeabilizing device in response to a desired value for said at least one parameter.
 10. The method of claim 1, wherein the step of transporting a substance across an outer surface of the area of membrane comprises: extracting a body fluid from or through said area of membrane; collecting said body fluid; and sensing the presence of said at least one analyte in said body fluid.
 11. The method of claim 1, wherein the substance is a drug.
 12. The method of claim 1, wherein the substance is a vaccine.
 13. The method of claim 1, wherein the substance includes at least one component of interstitial fluid.
 14. The method of claim 1, wherein the permeabilizing device is an ultrasound-producing device.
 15. The method of claim 1, wherein the permeabilizing device is a device that produces a force selected from the group consisting of chemical, electroporation, mechanical, disrupting, tape stripping, and laser forces.
 16. The method of claim 1, wherein the membrane is selected from the group consisting of biologic skin and synthetic skin.
 17. A method for enhancing a permeability of an area of skin comprising: increasing the permeability of the area of skin with a skin permeabilizing device; applying electricity to the area of skin; measuring at least one electrical parameter of the area of skin; and controlling the skin permeabilizing device based on the at least one electrical parameter.
 18. The method of claim 17, further comprising: applying electricity to the area of skin before increasing the permeability of the area of skin with a skin permeabilizing device; and measuring a baseline for the at least one electrical parameter.
 19. The method of claim 17, wherein the step of applying electricity to the area of skin comprises: applying a first source having a first frequency to the area of skin; and applying a second source having a second frequency to the area of skin.
 20. The method of claim 19, wherein the step of measuring a first electrical parameter comprises: measuring the at least one electrical parameter at the first frequency; and measuring the at least one electrical parameter at the second frequency.
 21. The method of claim 17, further comprising: coupling a first electrode with the area of skin; coupling a second electrode with the skin; and measuring the at least one electrical parameter using the first electrode and the second electrode.
 22. The method of claim 21, wherein the first electrode is coupled with a portion of stratum corneum of the area of skin.
 23. The method of claim 22, wherein the second electrode is coupled with a portion of epidermis over which the stratum corneum has been removed.
 24. The method of claim 22, wherein the second electrode is coupled with a portion of stratum corneum of the skin.
 25. The method of claim 21, wherein at least one of the first and second electrode is coupled through a conductive medium.
 26. The method of claim 25, wherein the conductive medium a gel.
 27. The method of claim 25, wherein the conductive medium is a liquid.
 28. The method of claim 21, wherein the second electrode is placed on the skin.
 29. The method of claim 17, further comprising: comparing the at least one electrical parameter with the baseline, and wherein the step of controlling the skin permeabilizing device comprises discontinuing the application of the skin permeabilizing device based on the comparison.
 30. The method of claim 19, wherein the first source comprises a voltage source having a frequency of about 10 Hz.
 31. The method of claim 19, wherein the second source comprises a voltage source having a frequency of about 1 kHz.
 32. The method of claim 17, wherein the parameter is selected from the group consisting of impedance, conductance, capacitance, current, and voltage.
 33. The method of claim 17, wherein said skin permeabilizing device comprises an ultrasound-producing device.
 34. The method of claim 20, wherein the step of controlling the skin permeabilizing device comprises decreasing a characteristic of the skin permeabilizing device when the measurement of the at least one electrical parameter at the first frequency and, the measurement of the at least one electrical parameter at the second frequency differ by less than a predetermined amount, the characteristic selected from the group consisting of intensity and duty cycle.
 35. The method of claim 19, wherein the step of controlling the skin permeabilizing device comprises decreasing a characteristic of the skin permeabilizing device when a rate of change between the one electrical parameters reaches a predetermined value, the characteristic selected from the group consisting of intensity and duty cycle.
 36. An apparatus for enhancing permeability of an area of skin comprising: a skin permeabilizing device configured to increase a permeability of the area of skin; an electrical source operable to apply electricity to the area of skin; a circuit to measure at least one electrical parameter of the area of skin; and a controller responsive to the circuit and operable to control the skin permeabilizing device.
 37. The apparatus of claim 36, wherein the electrical source comprises: a first source having a first frequency; and a second source having a second frequency.
 38. The apparatus of claim 37, wherein the circuit measures the at least one electrical parameter of the area of skin at the first frequency and to measure the at least one electrical parameter of the area of skin at the second frequency.
 39. The apparatus of claim 36, further comprising: a first electrode coupled on the area of skin; and a second electrode positioned on the skin; wherein the circuit measures the first parameter of the area of skin between the first electrode and the second electrode.
 40. The apparatus of claim 39, wherein the first electrode comprises an electrode that is coupled to a portion of stratum corneum of the area of skin.
 41. The apparatus of claim 40, wherein the second electrode comprises an electrode that is positioned on a portion of epidermis over which the stratum corneum has been removed.
 42. The apparatus of claim 39, wherein the first electrode comprises an electrode having a first surface area and coupled to a portion of stratum corneum of the area of skin.
 43. The apparatus of claim 42, wherein the second electrode comprises an electrode having a second surface area positioned on a portion of stratum corneum of the skin, and further wherein the second surface area is substantially larger than the first surface area.
 44. The apparatus of claim 40, wherein the controller compares the measurement of the at least one electrical parameter at the first frequency with the measurement of the at least one electrical parameter at the second frequency and turn off the skin permeabilizing device when they differ by less than about a predetermined amount.
 45. The apparatus of claim 37, wherein the first source comprises a voltage source having a frequency of about 10 Hz.
 46. The apparatus of claim 37, wherein the second source comprises a voltage source having a frequency of about 1 kHz.
 47. The apparatus of claim 38, wherein the parameter is selected from the group consisting of impedance, conductance, capacitance, current, and voltage.
 48. The apparatus of claim 37, wherein the controller compares the measurement of the at least one electrical parameter at the first frequency with the measurement of the at least one electrical parameter at the second frequency and decrease an intensity of the skin permeabilizing device source when they differ by less than about a predetermined amount.
 49. The apparatus of claim 38, wherein the controller compares the measurement of the at least one electrical parameter at the first frequency with the measurement of the at least one electrical parameter at the second frequency and decrease a duty cycle of the skin permeabilizing device source when they differ by less than about a predetermined amount.
 50. A method for enhancing permeability of an area of skin comprising: creating a volume of fluid adjacent the area of skin, said fluid having an initial concentration of a first substance; applying a skin permeabilizing device to the area of skin; monitoring changes in the concentration of the first substance; and controlling the skin permeabilizing device based on the changes in the concentration of the substance.
 51. The method of claim 50, wherein the volume of fluid is selected from the group consisting of a liquid, a gel, and a solid.
 52. The method of claim 50, wherein the first substance comprises an analyte.
 53. The method of claim 52, wherein the step of controlling the skin permeabilizing device comprises discontinuing the application of the skin permeabilizing device when the concentration of analyte in the volume of fluid increases to a predetermined concentration.
 54. The method of claim 50, wherein the step of controlling the skin permeabilizing device comprises discontinuing the application of the skin permeabilizing device when a rate of increase of analyte in the volume of fluid reaches a predetermined concentration.
 55. The method of claim 50, wherein the step of controlling the skin permeabilizing device comprises decreasing an characteristic of skin permeabilizing device when the concentration of analyte in the volume of fluid increases to a predetermined concentration, said characteristic selected from the group consisting of intensity and duty cycle.
 56. The method of claim 50, wherein the first substance comprises a benign substance and the initial concentration comprises a concentration higher than that found in the body.
 57. The method of claim 56, wherein the step of controlling the skin permeabilizing device comprises discontinuing the application of the skin permeabilizing device when the concentration of the benign molecule in the volume of fluid decreases to a predetermined concentration.
 58. The method of claim 56, wherein the step of controlling the skin permeabilizing device comprises discontinuing the application of the skin permeabilizing device when the rate of change in the concentration of the benign molecule in the volume of fluid reaches a predetermined value.
 59. The method of claim 56, wherein the step of controlling the skin permeabilizing device comprises decreasing an intensity of the skin permeabilizing device when the concentration of the benign molecule in the volume of fluid decreases to a predetermined concentration.
 60. The method of claim 56, wherein the step of controlling the skin permeabilizing device comprises decreasing an intensity of the skin permeabilizing device when the rate of change in the concentration of the benign molecule in the volume of fluid reaches a predetermined value.
 61. The method of claim 56, wherein the step of controlling the skin permeabilizing device comprises decreasing the intensity of a duty cycle of the skin permeabilizing device when the concentration of the benign molecule in the volume of fluid decreases to a predetermined concentration.
 62. A method for enhancing permeability of an area of skin comprising: creating a volume of fluid adjacent the area of skin; determining a reference value for an electrical parameter of the volume of fluid; applying a skin permeabilizing device to the area of skin; monitoring changes in the electrical parameter of the volume of fluid; and controlling the skin permeabilizing device based on the changes in the electrical parameter of the volume of fluid.
 63. The method of claim 62, wherein the volume of fluid is selected from the group consisting of a liquid, a gel, and a solid.
 64. The method of claim 62, wherein the electrical parameter is conductivity.
 65. A method for regulating the permeabilization of an area of skin, comprising: providing a first electrode in electrical contact with a first area of skin; providing a second electrode in electrical contact with a second area of skin; measuring an initial conductivity between said electrodes; applying a skin permeabilizing device to said first area of skin; measuring a second conductivity between said first and second electrodes; processing said initial conductivity and said second conductivity to establish information on a time-variation of skin conductance; calculating at least one parameter describing a kinetics of skin conductance changes responsive to said information; and terminating said application of said skin permeabilizing device in response to a desired value for said at least one parameter.
 66. The method of claim 65, wherein said application of said skin permeabilizing device is continuous.
 67. The method of claim 65, wherein said application of said skin permeabilizing device is discontinuous.
 68. The method of claim 65, wherein said skin permeabilizing device comprises an ultrasound-producing device.
 69. The method of claim 65, wherein said step of calculating at least one parameter describing a kinetics of skin conductance changes responsive to said information comprises: determining a slope of said information on a time-variation of skin conductance; and determining a point of inflection for said information on a time-variance of skin conductance.
 70. The method of claim 67, further comprising the steps of performing analog to digital conversion on said information on a time-variation of skin conductance; and filtering said digital data.
 71. The method of claim 65, wherein said step of processing said initial conductivity and said second conductivity to establish information on a time-variation of skin conductance comprises fitting said information into the following equation: $C = {{Ci} + \frac{\left( {C_{f} - C_{i}} \right)}{1 + ^{S{({t - t^{*}})}}}}$

where: C is current; C_(i) is current at t=0; C_(f) is the final current; S is a sensitivity constant; t* is the exposure time required to achieve an inflection point; and t is the time of exposure.
 72. A method for extraction and analysis of at least one analyte in a body fluid, comprising: increasing a permeability level of an area of skin; extracting a body fluid from or through said area of skin; collecting said body fluid; and sensing the presence of said at least one analyte in the body fluid.
 73. The method of claim 72, wherein said step of increasing a permeability level of an area of skin comprises applying ultrasound to said portion of skin.
 74. The method of claim 72, wherein said step of extracting a body fluid from or through said area of skin comprises applying a force selected from the group consisting of physical forces, chemical forces, biological forces, vacuum, electrical forces, osmotic forces, diffusion forces, electromagnetic forces, ultrasound forces, cavitation forces, mechanical forces, thermal forces, capillary forces, fluid circulation across the skin, electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces, acoustic forces, convective dispersion, photo acoustic forces, by rinsing body fluid off skin, and any combination thereof.
 75. The method of claim 74, wherein said vacuum force is applied continuously.
 76. The method of claim 74, wherein said vacuum force is applied discontinuously.
 77. The method of claim 74, wherein a material is placed between said vacuum force and said skin in order to maintain a surface configuration of said skin.
 78. The method of claim 77, wherein said material is selected from the group consisting of mesh, membrane, and perforated metal.
 79. The method of claim 77, wherein said vacuum force is generated by a device selected from the group consisting of mechanical, electro-mechanical, chemical, or electro-chemical.
 80. The method of claim 74, wherein said electrical force is selected from the group consisting of iontophoretic, electro-osmotic, and electroporation.
 81. The method of claim 74, wherein a gel is applied to said skin in order to encourage osmosis.
 82. The method of claim 74, wherein said ultrasound force is applied to create a result, said result selected from the group consisting of to pumping body fluid and fluid components, levitating, activating gas bodies, producing cyclic impulse mechanical stress to the skin, creating microstreaming, increasing temperature, and setting up standing waves.
 83. The method of claim 74, wherein a plurality of ultrasound-producing devices are used to create said ultrasound force.
 84. The method of claim 83 wherein said a plurality of ultrasound-producing devices have at least one different operating characteristic.
 85. The method of claim 84, wherein said operating characteristic is selected from the group consisting of frequency, intensity, and coupling media.
 86. The method of claim 74, wherein said mechanical forces are applied by a device selected from the group consisting of a roller, a squeezer, a stretcher, a compressor, and a tensioner.
 87. The method of claim 86, wherein said tensioner collects said body fluid in a cavity formed therein.
 88. The method of claim 74, wherein said thermal forces are created by a source selected from the group consisting of electric, chemical, ultrasonic, and optical energy sources.
 89. The method of claim 74, wherein temperature sensitive polymers are used to extract body fluids.
 90. The method of claim 72, wherein said step of collecting said body fluid comprises using a collection method selected from the group consisting of absorption, adsorption, phase separation, mechanical, electrical, chemically induced, capillary forces, and a combination thereof.
 91. The method of claim 90, wherein said absorption collection method comprises collecting said body fluid into a gel.
 92. The method of claim 91, wherein said gel contains a captive enzyme.
 93. The method of claim 90, wherein said phase separation method comprises isolating said body fluid with an appropriate density immiscible fluid.
 94. The method of claim 93, further comprising collecting said body fluid into a conical chamber.
 95. The method of claim 90 wherein a hydrophobic coating is applied to said skin prior to said step of extracting a body fluid from said area of skin.
 96. The method of claim 75, wherein said body fluid is collected from said hydrophobic coating.
 97. The method of claim 90, wherein said mechanical collection method comprises applying a force selected from the group consisting of vacuum, pressure, and acoustic forces.
 98. The method of claim 90, wherein said electrical collection method comprises moving a charged object from said skin to a collecting compartment using electrical forces.
 99. The method of claim 90, wherein said chemical collection method comprises applying a hydrophilic gel to collect body fluids.
 100. The method of claim 90, wherein said capillary collection method comprises: filling at least one capillary with a plurality of fibers; and collecting said body fluid in said at least one capillary.
 101. The method of claim 72, wherein said step of sensing the presence of at least one analyte comprises applying a sensing method selected from the group consisting of electrochemical, optical, acoustical, biological, enzymatic technology, and combinations thereof.
 102. The method of claim 72, wherein living cells are used to sense a concentration of an analyte in body fluid.
 103. The method of claim 72, further comprising the step of providing an output for a user interface comprises providing an alarm that indicates an abnormal analyte concentration.
 104. The method of claim 72, further comprising the step of providing an output for a user interface comprises providing an trend information.
 105. The method of claim 72, further comprising the step of providing history information.
 106. The method of claim 72, wherein said user output is downloadable.
 107. A system for extraction and analysis of at least one analyte in a body fluid comprising: a transducer for increasing the permeability of an area of skin; an extraction device for extracting interstitial fluid from said area of skin; a collection device for collecting said extracted interstitial fluid; and a sensing device for sensing the presence of at least one analyte in said extracted interstitial fluid.
 108. The system of claim 107, further comprising a microcontroller for controlling at least one of said transducer, said extraction device, said collection device, and said sensing device.
 109. The system of claim 107, further comprising a user output device.
 110. The system of claim 108, further comprising a microcontroller for controlling said user output device.
 111. The system of claim 107, wherein said transducer comprises an ultrasonic transducer.
 112. The system of claim 107, wherein said extraction device is a device that produces a force selected from the group consisting of physical forces, chemical forces, biological forces, vacuum pressure, electrical forces, osmotic forces, diffusion forces, electromagnetic forces, ultrasound forces, cavitation forces, mechanical forces, thermal forces, capillary forces, fluid circulation across the skin, electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces, acoustic forces, convective dispersion, photo acoustic forces, by rinsing body fluid off skin, and any combination thereof.
 113. The system of claim 107, wherein said collection device is a device that uses a collection method selected from the group consisting of absorption, adsorption, phase separation, mechanical, electrical, chemically induced, and a combination thereof.
 114. The system of claim 107, wherein said sensing device is a device that senses the presence of an analyte by a sensing method selected from the group consisting of electrochemical, optical, acoustical, biological, enzymatic technology, and combinations thereof.
 115. The system of claim 109, wherein said user output device provides information selected from the group consisting of trend information, history information, operating information, and combinations thereof.
 116. The system of claim 115, wherein information from said user output device is downloadable to a computer.
 117. A method for blood glucose determination comprising: increasing a permeability of an area of skin; extracting interstitial fluid from said area of skin; collecting said interstitial fluid in a gel, said gel containing at least one glucose sensitive reagent that changes at least one characteristic of said gel when glucose is present; and monitoring a change in said at least one characteristic of said gel.
 118. A system for blood glucose determination comprising: a transducer for increasing the permeability of an area of skin; an extraction device for extracting interstitial fluid from said area of skin; a collection device for collecting said extracted interstitial fluid; a gel in said collection device; at least one glucose sensitive reagent that changes at least one characteristic of said gel when glucose is present; and a monitoring device for monitoring a change in said at least one characteristic of said gel.
 119. The system of claim 118, wherein the at least one glucose sensitive reagent is in said gel.
 120. A drug delivery patch apparatus, comprising: a transducer for applying ultrasound to a membrane; a power source coupled to said transducer; a plurality of drug molecules between said transducer and said biological membrane; an attaching device for attaching said drug molecules and said transducer to said membrane; and a user interface that interacts with said transducer, said power source and said drug molecules.
 121. The drug delivery patch apparatus of claim 120, further comprising drive electronics coupled to said transducer, said drive electronics enables said transducer to apply ultrasound.
 122. The drug delivery patch apparatus of claim 120, wherein said membrane is skin.
 123. The drug delivery patch apparatus of claim 120, wherein said drug molecules and said attaching device are combined.
 124. The drug delivery patch apparatus of claim 120, wherein said drug molecules are suspended in the group consisting of a liquid, a gel, and a solid matrix.
 125. The drug delivery patch apparatus of claim 120, wherein said power source is coupled to said transducer via hardwire.
 126. The drug delivery patch apparatus of claim 120, wherein said power source is coupled to said transducer via telemetry.
 127. The drug delivery patch apparatus of claim 120, wherein said transducer comprises acoustic elements, such that said acoustic elements are swept in a temporal nature as said transducer applies ultrasound.
 128. The drug delivery patch apparatus of claim 120, wherein said attaching device includes a band for attaching to said membrane.
 129. The drug delivery patch apparatus of claim 120, further comprising a feedback mechanism coupled to said drug molecules and said user interface for monitoring the amount of said drug molecules.
 130. The drug delivery patch apparatus of claim 120, further comprising a feedback mechanism coupled to said transducer and said user interface for monitoring the amount of ultrasound applied to said membrane.
 131. The drug delivery patch apparatus of claim 120, wherein said user interface is coupled to said transducer, said power source and said drug molecules by telemetry.
 132. The drug delivery patch apparatus of claim 120, wherein said user interface is coupled to said transducer, said power source and said drug molecules by an infrared device.
 133. The drug delivery patch apparatus of claim 120, wherein said user interface is coupled to said transducer, said power source and said drug molecules by fiber optics.
 134. The drug delivery patch apparatus of claim 120, wherein said transducer is configured as a cylinder.
 135. The drug delivery patch apparatus of claim 120, wherein said transducer is configured as a hollow cylinder.
 136. The drug delivery patch apparatus of claim 120, wherein said transducer is a hemispherical configuration.
 137. The drug delivery patch apparatus of claim 120, wherein said transducer is a conical configuration.
 138. The drug delivery patch apparatus of claim 120, wherein said transducer is a planar configuration.
 139. The drug delivery patch apparatus of claim 120, wherein said transducer is a rectangular configuration.
 140. The drug delivery patch apparatus of claim 120, further comprising a device for creating a driving force that operates in conjunction with said transducer, said driving force selected from the group consisting of physical forces, chemical forces, biological forces, vacuum pressure, electrical forces, osmotic forces, diffusion forces, electromagnetic forces, ultrasound forces, cavitation forces, mechanical forces, thermal forces, capillary forces, fluid circulation across the skin, electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces, acoustic forces, convective dispersion, photo-acoustic forces, by rinsing body fluid off skin, and any combination thereof.
 141. The drug delivery patch apparatus of claim 120, further comprising a limiting step membrane between said membrane and said drug molecules.
 142. The drug delivery patch apparatus of claim 120, wherein said membrane is pretreated by a force.
 143. The drug delivery patch apparatus of claim 142, wherein said force is selected from the group consisting of physical forces, chemical forces, biological forces, vacuum pressure, electrical forces, osmotic forces, diffusion forces, electromagnetic forces, ultrasound forces, cavitation forces, mechanical forces, thermal forces, capillary forces, fluid circulation across the skin, electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces, acoustic forces, convective dispersion, photo-acoustic forces, by rinsing body fluid off skin, and any combination thereof.
 144. A method for delivering a drug through a membrane, the method comprising the steps of: supplying power to a transducer that applies ultrasound; delivering drug molecules through said membrane by applying ultrasound to said drug molecules and having an attaching device for attaching at least said drug molecules and said transducer to said membrane.
 145. The method of claim 144, further comprising the step of: pre-treating said membrane by applying a force prior to said delivering step, said force selected from the group consisting of physical forces, chemical forces, biological forces, vacuum pressure, electrical forces, osmotic forces, diffusion forces, electromagnetic forces, ultrasound forces, cavitation forces, mechanical forces, thermal forces, capillary forces, fluid circulation across the skin, electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces, acoustic forces, convective dispersion, photo-acoustic forces, by rinsing body fluid off skin, and any combination thereof.
 146. The method of claim 144, wherein said delivering step includes applying a driving force in conjunction with said ultrasound from said transducer, said driving force selected from the group consisting of physical forces, chemical forces, biological forces, vacuum pressure, electrical forces, osmotic forces, diffusion forces, electromagnetic forces, ultrasound forces, cavitation forces, mechanical forces, thermal forces, capillary forces, fluid circulation across the skin, electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces, acoustic forces, convective dispersion, photo-acoustic forces, by rinsing body fluid off skin, and any combination thereof.
 147. A method for transdermal vaccination comprising: increasing permeability of an area of skin; providing a vaccine on said area of skin; delivering said vaccine to at least one skin cell.
 148. The method of claim 147, wherein said step of increasing a permeability of an area of skin comprises applying ultrasound to said area of skin.
 149. The method of claim 147, wherein said step of providing a vaccine on said area of skin comprises providing a patch containing said vaccine on said area of skin.
 150. The method of claim 147, wherein said step of providing a vaccine on said area of skin comprises providing a gel containing said vaccine on said area of skin.
 151. The method of claim 147, wherein said step of providing a vaccine on said area of skin comprises providing a liquid containing said vaccine on said area of skin.
 152. The method of claim 147, wherein said step of delivering said vaccine to at least one skin cell comprises diffusing said vaccine to said at least one skin cell.
 153. The method of claim 147, wherein said steps of increasing a permeability of a area of skin and providing a vaccine on said area of skin are simultaneous.
 154. The method of claim 147, wherein said vaccine is selected from the group consisting of a peptide, a protein, DNA, an allergen, and other antigens.
 155. The method of claim 147, wherein said at least one skin cell is selected from the group consisting of Langerhans cells, dendric cells, and keratinocytes.
 156. The method of claim 147, wherein said step of delivery said vaccine to said immune cell comprises applying a driving force selected from the group consisting of physical forces, chemical forces, biological forces, vacuum, electrical forces, osmotic forces, diffusion forces, electromagnetic forces, ultrasound forces, cavitation forces, mechanical forces, thermal forces, capillary forces, fluid circulation across the skin, electro-acoustic forces, magnetic forces, magneto-hydrodynamic forces, acoustic forces, convective dispersion, photo acoustic forces, and any combination thereof.
 157. A method for transdermal vaccination and immunization, comprising: applying ultrasound to irritate a area of skin; and providing a vaccine to said area of skin. 